QUESTIONS FOR EXAMINATION ON BIOCHEMISTRY
MODULE I. STRUCTURE, FUNCTION AND PROPERTIES OF PROTEINS. ENZYMES
1. The functions of proteins in the human body. Physiologically important peptides.
Levels of protein structure. Primary, secondary, tertiary, quaternary structures.
Chemical bonds, providing their stability. Globular, fibrillar, transmembrane proteins:
structural features, examples. Protein folding. Chaperonins. Protein denaturation and
renativation. Prion disease.
Functions of proteins:
- Proteins take part in chemical reactions as catalysts
- Formation of cell membrane, extracellular and intracellular components of cells
- Regulation of metabolism, gene expression, and physiologic function of visceral organs
(hormones)
- Defence function of organism against foreign antigens from environmental factors
- Transport of low molecular weight substance into/backward the cell
Physiological important peptides: natriuretic peptide (2 types: atrial NP and brain NP)
- Increase sodium secretion in kidney, reduce extracellular fluid, decrease blood
pressure.
Levels of protein structure:
Primary: covalent backbone of polypeptide chain formed by specific sequence of amino
acids residues connected to each other by peptide bond. Primary structure has pivotal
role in formation higher level of protein structures.
Secondary: polypeptide chain conformation in 3-dimensional structure resulted from
interactions between neighbour aa residues by hydrogen bonds.
Tertiary: refers to folding polypeptide chain. This structure is stabilized by hydrogen
bond, Van der Waals or disulfide bond.
Quaternary: forming bonds between different polypeptide chains by noncovalent
bonds (van der Waals, electrostatic, hydrophobic) and covalent difulfide bridges. (Eg:
hexokinase: 2 polypeptide chains, RBC: 4 polypeptide chains)
Globular, fibrillar, transmembrane proteins:
Globular proteins: globular shape. Eg: albumin, globulins
Fibirillar proteiins: long fibrous shape. Eg: collagen, elastin
Transmembrane proteins: located in membrane and connect with both extracellular
and intracellular matrix. Eg: GLUT, Na+/K+ ATPase, etc
Protein folding: most of polypeptide chains undergo folding to form the most stable
conformation to function. The folding process takes place immediately after synthesis in
intracellular matrix and facilitated by chaperones, chaperonins.
- Chaperones: facilitate forming quaternary structure and prevent denaturation of
polypeptide chain
- Chaperonins: conformational change of protein coupled with hydrolysis of ATP and
many different enzymes
Protein denaturation and reactivation:
Protein denaturation: the loss of native conformation and function of proteins
- Factors: heat, pH, organic solvents (alcohol, acetone), urea, etc
- Those factors destroy weak bonds: hydrophobic, electrostatic bonds.
- Denatured enzymes lose their catalytic properties.
- Denaturation is reversible. The process regain protein conformation is called
renaturation. Eg: ribonuclease (inactivated by urea and reactive by remove urea)
Prion disease: Creutzdelt-Jakob disease and Kuru
Prion are proteins which have role in cell-cell communications and intracellular
signaling.
Causes:
- Emerging prion gene mutation
- Contagious way: prion penetrating human body with meal. It arises prion disease in
human when it comes from malady prion diseased animals.
- Sporadic.
2. Complex proteins. Classification, examples. Structure and functions of myoglobin.
Complex protein: 2 parts: protein and non-protein parts
Classification:
Glycoprotein
Include carbohydrates
Lipoprotein
Nucleoprotein
Phosphoprotein
Include lipids
Include nucleic acids
Include residues
phosphoric acid
Include metals
Metaloprotein
Hyaluronic acid, keratan sulfate, dermatan
sulfate,…
Palmitic acid, muristic acid
Nucleohistone, ADP, GMP, NADP, etc
Casein (milk), ovalbumin (egg), vitellins (egg
yolk)
Hemoprotein (Fe): hemoglobin, ferritin
Copper protein (Cu): cytochrome c oxidase
Zn protein: RNA polymerase, DNA polymerase
Mo, Co, Mn containing proteins
Besides, we can have different classifications: chromoprotein (color proteins) and
flavoproteins (conjugated protein containing flavin group)
Myoglobin:
- Structure: composed of 1 coiled polypeptide globin and a heme group. The heme
consists of a ferrous (Fe2+) in the center of porphyrin ring.
- Function: myoglobin can bind to oxygen stronger than Hg so it function as storage of
oxygen in skeletal muscles.
3. The structure and functions of hemoglobin. T- and R-forms of hemoglobin. Allosteric
effects of hemoglobin: cooperative effect, Bohr effect, effect of 2,3-diphosphoglycerate.
Features of fetal hemoglobin. The concept of hemoglobinopathies. Sickle-cell anemia.
Thalassemia.
Hemoglobin
- Structure: has biconcave shape, comprises of four subunits, each having one
polypeptide chain and one heme group. 4 polypeptide chains forming globin group.
Heme group is composed of a ringlike organic compound known as a porphyrin to which
an iron atom is attached.
- Function: take up and release gases: oxygen and CO2
- T form: deoxyhemoglobin state of Hb
- R form: fully oxygenated state of Hb
Allosteric effects of hemoglobin:
- Cooperative effect: binding of the 1st oxygen to Hb increase affinity of Hb to the 2nd, 3rd
and then 4th O2
- Bohr effect: binding of CO2 forces the release of O2 from Hb
- 2,3-diphosphoglycerate: produced from 1,3DPG - a glycolytic intermediate. 2,3DPG
normally bind and stabilize T form of Hb. When T form is oxygenated into R
conformation, 2,3 DPG is ejected.
Fetal hemoglobin: HbF
- Contain 4 chains: 2 alpha and 2 gamma globin chains
- Have stronger affinity to oxygen compare to HbA due to gamma globin. - 2,3 DPG is
unable bind to HbF due to gamma chain
- Amount of HbF gradually decreases with growing of newborn
Hemoglobinopathies: mutations of globin genes. 2 types
Sickle cell anemia: qualitative abnormalities
Thalassemia: quantitative abnormalities
Sickle cell anemia:
- Substitution of Valine for glutamic acid at 6th position in beta chain of HbA
- When HbS is reduced (in low O2 tension) precipitate into crystals in RBC changes shape
become sickle shaped and reduced Hb ability to bind with O2
- Effects: blockage of microcirculation, increases blood viscosity, hemolysis, anemia.
- Treatment: bone marrow transplantation, drugs: formation of HbF which decreases
polymerization of deoxygenated Hb by Azacytidine and hydroxyurea.
Thalassemia: inherited disease which cause hypoproduction of hemoglobin chain
- Alpha thalassemia: reduced in production of alpha chains of hemoglobin
- Beta thalassemia: reduced in production of beta chains of hemoglobin (common)
Risk factors: family history of thalassemia, certain ancestry (from African Americans or
Southeast Asian)
Complications: iron overload, enlarged spleen, slow growth rate, heart problems, bone
deformities, dark urine, yellow or pale skin
Diagnosis: blood test: test for anemia and abnormal hemoglobin
Treatment: blood transfusion, bone marrow transplantation, medications, spleen or
gallbladder removal surgery.
4. General characteristics of enzymes. Enzyme specificity: types, examples. The structure
of enzymes. Cofactors and coenzymes. Basic reaction types and classes of enzymes.
Systematic names of enzymes.
Enzyme: catalyst of chemical reactions in human body
- Enzymes are not consumed during chemical reactions
- They don’t cause the reactions; they ONLY speed up the rate of reaction
- Enzymes have high specificity for the reaction it catalyze
- Enzyme made up from proteins, it can function in normal condition (pH, temperature)
- Enzymes possess high catalytic power and their catalytic power can be regulated.
Enzyme specificity:
- Absolute specificity: enzyme catalyzes for chemical reaction of only one single
substrate
- Relative specificity: enzyme can catalyze for chemical reactions of some substrates
with identical structure or functional group
- Stereospecificity: enzyme can catalyze for chemical reaction of certain stereoisomers
(L or D) of substrate
Structure of enzyme: simple or complex proteins. APO + COE = HOLOENZYME
- Apoenzyme (protein part): made up from polypeptide, sensitively damaged by pH,
heat
- Coenzyme (nonprotein part): not sensitive to damage of temperature, pH. Eg: watersoluble vitamin: NAD, FAD, FMN
+ Cofactor: additional chemical compound for enzyme function: inorganic ion: Fe, Cu,
Mg
Classification: enzymes are classified due to the reaction it catalyzed: 6 main groups:
1. Oxidoreductase: oxidation/reduction reaction.
2. Transferase: transferation of functional group between donor and acceptor
substrates
3. Hydroxylase: hydrolysis of substrates
4. Lyases: non-hydrolytic destroy of substrates
5. Isomerase: intramolecular transferation of functional group
6. Ligases (synthetases): addition to substrate to form new product
Systematic name of enzyme: consists of 2 part: substrate name – type of reaction
enzyme catalyze.
5. Catalytic mechanisms of enzymes. Active catalytic site. Stages of enzymatic reaction.
Models of interaction of enzyme with its substrate.
Catalytic mechanism of enzymes:
Normally, conversion of substrate S to product P need energy (delta G). Enzyme
transforms the substrate to transition state S* which needs less free energy to be
converted to P. The amount of free energy needed to transform substrate S to transition
state S* is called activation energy.
E + S <-> ES <-> ES* <-> P+ E
Active catalytic site: is where substrate can bind complementary with enzyme. The
binding of substrate and enzyme mediated by multiple weak bonds (hydrogen,
hydrophobic, electrostatic, van der Waals interactions).
There are 2 types of recognition between enzyme and substrate:
- Lock and key: substrate perfectly fit active site in enzyme
- Induced fit: substrate comes to active site and enzyme changes its active site fit to
substrate.
Stages of enzymatic reaction: E + S <-> ES <-> ES* <-> P+ E
- Forming enzyme-substrate complex
- Brings substrate to its transition state
- Forming product. Product and enzyme release each other.
- Enzyme continues forming new ES complex
Models of interaction of enzyme with its substrate:
1. Proximity: enzyme brings 2 molecules together in solution
2. Orientation: Substrates don’t form products even with enough energy. Enzymes
bind to substrates to orient reactive groups are steered to the direction can lead
to product.
3. Deformation of molecule of substrate: enzyme forces substrate undergo
cleavage
4. Acid-base catalysis: exchange protons (from aa) between weak organic acids and
bases in active site
5. Covalent catalysis: forming transient covalent bond between enzyme and
substrate for activation of substrate for further reaction
6. Metal ion catalysis: ion interaction between metal enzyme and substrate help
orient the substrate for reaction to happen or mediate oxidation-reduction
reaction.
6. Enzyme kinetics. The dependence of enzyme reaction rate on substrate
concentration. The Michaelis-Menten equation. The dependence of enzyme reaction
rate on amount of enzyme. The dependence of enzyme reaction rate on pH and
temperature.
Enzyme kinetics: interaction between velocity of reaction and concentration of
substrate
Ability to catalyze chemical reaction of enzyme is
saturated (show by decreasing in catalyze rate) with
the increasing of substrate concentration.
- When [S] is low: rate ­­concentration
- When [S] is high, rate is constant and independent
to [S]
Michaelis-Menten equation: is substrate
concentration at which velocity is half maximum
Dependence of enzyme reaction on amount of enzyme: directly proportional because
there are more active binding site available for substrates.
Enzyme basically proteins. It depends on pH and temperature.
- All proteins can function well with range of temperature from 4 to 57°C and optimal
temperature is 37°C.
- Different enzyme in different condition of different organs require different range of
pH. Eg: G6P pH ranges from 6-10; pepsin pH ranges from 0 to 6.
7. Inhibition of enzyme activity. Competitive and noncompetitive inhibition. Irreversible
inhibition. Enzyme kinetic changes. Examples.
Inhibition of enzyme: interfere the catalysis of enzyme and slowing or stopping the
reaction rate
2 types: reversible and irreversible
Reversible: there is dissociation between enzyme-inhibitor complex
1. Competitive: inhibitors compete with substrate for active binding site
Eg: succinate and malonate both have carboxyl group at 2 terminal ends. Both
compete for succinate dehydrogenase (in TCA)
2. Non-competitive: inhibitors bind to substrate or substrate-enzyme complex in
distinct site to active site then stop the reaction to occur.
3. Uncompetitive: inhibitors bind to substrate enzyme complex at distinct site from
active site
Irreversible: inhibitors dissociate from enzymes slower because it forms tight bond to
enzyme. Eg: enzyme contains serine residue can be irreversible inhibit by DPP or
enzyme contains cysteine residue can be irreversible inhibit by iodoacetamide
Enzyme kinetic changes:
Ability to catalyze chemical reaction of enzyme is
saturated (show by decreasing in catalyze rate) with
the increasing of substrate concentration.
- When [S] is low: rate ­­concentration
- When [S] is high, rate is constant and independent
to [S]
8. Allosteric enzymes. Structure and functions. Allosteric effectors. Regulation of
enzyme activity by protein-protein interactions. Regulatory proteins. Examples.
Regulation of enzyme activity by phosphorylation and dephosphorylation and by partial
proteolytic cleavage.
Allosteric enzymes
- Structure: enzyme contains 2 binding sites: active site for substrate and allosteric site
for effector.
- Function: their conformation, affinity to substrate can be change by the binding of
effector to allosteric site.
Allosteric effectors: 2 types of classification
- Activator: activate catalytic function of
enzyme
- Heterotropic: effector differs to
substrate
- Inhibitor: inhibit catalytic function of
enzyme
- Homotropic: effector and substrate are
the same
Regulation of enzyme activity by protein-protein interactions
- Protein (eg: neurotransmitter) and protein receptor interaction brings to
conformational change and then biochemical events that activate or inhibit the activity
of the enzyme
Eg:
Regulation of enzyme activity by phosphorylation and dephosphorylation
- Phosphorylation and dephosphorylation can bring to activate or inhibit of enzyme
Eg: ATP (active) -dephosphorylation-> ADP (inactive) + Pi
Glycogen synthetase (active) -phosphorylation-> (inactive)
Regulation of enzyme activity by partial proteolytic cleavage.
- Proenzyme contains active enzyme and inhibit polypetide by protases. The cleavage of
inhibit polypeptide activates active enzyme
9. Isozymes. Examples. Biologic role. Enzymopathy. Examples. Enzymodiagnostics and
enzymotherapy. Examples of using enzymes in therapy. Abzymes.
Isozymes: proteins with different structure, different polypeptide chain, different gene
coding, different localization (tissues, organs) but catalyse the same reaction.
Eg:
Glucose -> glucose 6 phosphate. Hexokinase (muscle), glucose kinase (liver)
Creatine kinase BB (brain and smooth muscle) , MB (cardiac muscle), MM (
skeletal muscle)
Biological role:
- Different tissues in different organs with different condition (pH, etc) all require the
same reaction (for metabolism) need different enzymes with suitable optimal
conditions.
Enzymopathy: hereditary disease in inborn that results in enzyme disorder
Eg: Abnomalities in aa metabolism by alkaptonuria or albinism
Abnormalities in lipid metabolism by Tay Sach disease or Niemann-Pick disease
Abnormalities in carbohydrate metabolism maybe manifested by diabetes mellitus or
galactosemia
Enzymodiagnostic: enzyme can be used as biomarkers for many diseases: pancreatitis
(amylase, lipase), myocardial infarction (lactate DH), liver disease (AST, ALT)
Enzymotherapy: using enzyme for replacement treatment in deficiency or absence of
necessary enzymes.
Trypsin
Ribonuclease
Treatment of wounds, bronchititis, pneumonia, tuberculosis
Treatment of wounds, bronchitis, pneumonia, encephalitis, tuberculosis
Collagenase
a-amylase
Thrombin
Treatment of wounds
Pancreatitis, hepatitis
Bleeding
Abzymes: is an antibody that is used to lower the activation energy of transition stage in
catalysation. Abzymes are typically artificial made.
MODULE II. MEMBRANES. BIOLOGIC OXIDATION.
1. Structure of biological membranes. Membrane lipids: structure, function. Membrane
proteins. Properties of biological membranes: microviscosity, asymmetry, fluidity. Fluid
mosaic model of membrane structure.
Structure of biological membrane
- Sheet like structure and noncovalent assemblies. It closes boundaries between
different intracellular compartments
- Main compounds are phospholipid and protein. They held together by noncovalent
interactions.
- Membrane composed of hydrophilic (glycerol head) and hydrophobic (fatty acid tail)
moieties. Phospholipids form bilayer structure which is a barrier for polar molecules.
- Membranes have asymmetric structure: outside and inside surfaces differ from each
other
- Membrane potential is important in transport, excitability, and energy conversion
Membrane lipid:
- Main components: phospholipids, glycolipids, and cholesterols.
- Structure: lipid bilayer consists of 2 leaflets: outer and inner and they are different in
content of phospholipids; fluid structure
- Function: barrier control entrance and exit of ions, substances, impermeable to water
and water-soluble substances; different types of lipids make up and provide lipid
membrane characteristics (stiffness, flexibility, rigid, etc); endocytosis/exocytosis
Membrane proteins: serve as receptors, enzymes, pumps, channels, energy transducers
for all transportation, signalling activities of the cell
- Peripheral: bond to membrane by weak interactions
- Integral: imbedded to lipid bilayer by interact w hydrocarbon chain of membrane
Properties of biological membrane:
- Microviscosity: allow diffusion of substances down its electrochemical gradient
- Asymmetric: component and structure different between inner and outer leaflets
- Fluidity: lipids provide a fluid structure because lipid molecule is capable of lateral
diffusion
Fluid mosaic model of membrane structure: floating proteins in phospholipid molecules
- Lipid bilayer has dual role: a solvent for membrane and a barrier of permeability
- Some membrane phospholipids are linked by membrane proteins
- Membrane proteins undergo lateral diffusion
2. Transport of substances through biological membranes. Passive transport. Ion
channels. Aquaporins. Active transport of substances across membranes. Primary active
transport. Structure of Na+, K+-ATP-ase. Secondary active transport. Biochemical
mechanisms of endocytosis and exocytosis.
Transport of substances through biological membranes: 2 types
• Passive: not require energy, down concentration gradient
- Simple diffusion: movement of molecules (gases) across the membrane
- Facilitated diffusion: movement of hydrophilic molecules (water, glucose, amino
acids, ions: Na+, K+, Ca2+, Cl-) by channels, carrier proteins which are integral
proteins
+ Aquaporins: channels facilitate water transport down gradient
• Active: transfer substances against concentration gradient coupled with
hydrolysis of ATP
- Primary active transport: directly using ATP as energy for transport of substrates
(Na+/K+ ATPase; H+ ATPase; Ca2+ ATPase)
- Secondary active transport: indirectly using ATP for transport of substrates
(Glucose Na+). There are 2 types:
+ Uniport: simple diffusion of transported molecule through membrane towards
its lower concentration
+ Cotransport/antiport: transport of substance across membrane with
simultaneous transport of another substance (Glucose-Na+ into the cell)
Endocytosis and exocytosis: transport of macromolecules (proteins, nucleic acids, and
polysaccharides
Endocytosis: engulfing of macromolecules by the cells from the external environment.
Invagination to the membrane, vesicle gradually form and
2 types of endocytosis:
- Phagocytosis: uptake large solid particles (bacteria, viruses, cell fragments) and
performed by phagocytes
- Pinocytosis: process of absorption of liquids containing dissolved substances
(glucose, polysaccharides); all cells are capable of pinocytosis
Exocytosis: segregation of macromolecules from inside to outside the cell
Cell synthesizes macromolecules (fibroblast secretes collagen, beta cell secrete insulin,
glandulocyte of stomach secretes zymogens). Macromolecule moves into protrusion
formed on the cell membrane then transforms into vesicle and enter extracellular
medium.
3. Transmembrane signal transduction. Membrane receptor proteins. G-proteins:
structure, function, regulation.
Transmembrane signal transduction: the process after binding to extracellular
substance, integral protein sends physical or chemical signal and stimulates series of
intracellular reactions
Membrane receptor proteins: integral proteins which have 3 dimensional structure
- Made up from glycoprotein or lipoprotein, etc
- 3 types: ion channel-linked receptors, G-protein-linked receptors, and enzyme-linked
receptors
G proteins (guanine nucleotide-binding proteins):
- 2 types: monomeric (enzyme in cytosol) and heterotrimetric (involved in signal
transduction)
- Structure of heterotrimetric: composed of alpha, beta, and gamma subunits
- Function: activated G protein-coupled receptor stimulates G protein to activate further
cascade of intracellular reactions which finally leads to cell function changes.
- Regulation: binding between hormones, neurotransmitters, or other signalling factors
to receptors
4. Catabolism and anabolism. The scheme of metabolism and energy transfer in human
organism. Specific and common catabolic pathways.
Catabolism and anabolism are 2 stages of metabolism:
- Anabolism: combining building blocks compounds into macromolecules which
required by organism. This process require energy. End products are proteins, nucleic
acids, polysaccharides.
- Catabolism: breaking down of complex molecules to simpler ones accompanied with
releasing energy. The end products are inorganic substances such as CO2, water.
Specific and common catabolic pathway:
Catabolism consists of 3 stages:
- I stage: macromolecule (proteins, nucleic acids, fats) breaking down to building blocks
(glucose, glycerol, aa, fatty acids, etc) SPECIFIC
- II stage: conversion of I stage product to common metabolites (pyruvate, acetyl CoA)
COMMON
- III stage: create ATP from NADH, FADH2 and FMNH2 through citric acid cycle, terminal
oxidation, oxidative phosphorylation COMMON
5. Oxidative decarboxylation of pyruvate. Structure of the pyruvate dehydrogenase
complex. Stages of the pyruvate oxidative decarboxylation. Regulation of the pyruvate
dehydrogenase complex.
Oxidative decarboxylation of pyruvate: transition of pyruvate to acetyl CoA under
catalization of pyruvate dehydrogenase complex
Pyruvate dehydrogenase complex: located in inner mitochondria membrane, consist of
3 enzymes
- Pyruvate dehydrogenase (E1) (cofactor TPP)
- Dihydrolipoyl transacetylase (E2)
- Dihydrolipoyl dehydrogenase (E3)
- E3 binding protein
Stages of pyruvate oxidative decarboxylation:
- I: decarboxylation of pyruvate with cofactor TPP (B1)
- II: transfer of acyl group to CoA SH to produce Acetyl CoA
- III: reducing NAD+ forming NADH and ATP
Regulation of pyruvate DHC: (on-off by allosteric mechanism; hormone)
+ Activation: Insulin, Ca2+, Mg2+, NAD+, CoA, Pyruvate
+ Inhibition: ATP, NADH, Acetyl CoA, arsenite (inhibit cofactor).
6. Tricarboxylic acid cycle. Overview, energetics of the tricarboxylic acid cycle.
TCA cycle is a series of enzyme catalyzed the common pathway of catabolism create
energy. Acetyl CoA which is product of carbohydrates, fatty acids, ketone bodies and aa
catabolism enters the cycle.
TCA cycle can generate 3 NADH and 1 FADH2 from 1 Acetyl CoA
7. Reactions of the tricarboxylic acid cycle. Reactions catalyzed by dehydrogenases.
Reaction of the substrate-level phosphorylation.
Reactions catalyzed by DH
Isocitrate ----- isocitrate DH ----> alpha ketoglutarate
Alpha ketoglutarate ------ alpha ketoglutarate DH ---> succinyl CoA
Succinate ------ succinate DH ------> fumarate
Malate ------- malate DH ------> oxaloacetate
Reactions of substrate level phosphorylation
8. Regulation of the tricarboxylic acid cycle. Anabolic functions of TCA cycle. Anaplerotic
reactions.
Regulation of TCA cycle is controlled at several points
• OAA + Acetyl CoA ---> Citrate
- Inhibitors: ATP, NADH, Succinyl CoA
• Isocitrate ----> alpha ketoglutarate
- Inhibitors: ATP, NADH
- Activators: ADP, Ca2+
• Alpha ketoglutarate ----> succinyl CoA
- Inhibitors: NADP, succinyl CoA, ATP
Anabolic functions of TCA cycle: intermediates of TCA cycle are used as precursors in
biosynthesis of many compounds (carbohydrates, lipids, heme, amino acids)
Anaplerotic reactions: chemical reactions that form intermediates of TCA cycle
- Pyruvate ----- pyruvate carboxylase ----> oxaloacetate (gluconeogenesis)
- Glutamate ------- glutamate DH ------> alpha ketoglutarate (deamination)
- Fatty acids ------> succinyl CoA (beta oxidation of fatty acid)
- Adenylsuccinate ----- adenylosuccinate lyase ---à fumarate (purine synthesis)
9. Macroergic substrates. Classification of high-energy bond containing substances. ATP
as universal “energetic currency”.
Macroergic substrates are high energy storage compounds, also known as high-energy
phosphate compounds, chiefly found in muscular tissue in animals. A macroergic
substance is a substance able to release at least 25 kJ of energy per mol.
Classification of high-energy bond containing substances: 5 groups
1- Pyrophosphates: contain two phosphorus atoms in a P-O-P linkage (ATP)
2- Enol phosphates: is formed when phosphate group attaches to a hydroxyl group which
is bounded to a carbon atom having double bond (Phospho Enol Pyruvate)
3- Acyl phosphates: is formed by the reaction between carboxylic acid group and
phosphate group. (1,3 biphospho-glycarate carbonyl phosphate)
4- Thiol phosphates: results from the reaction between thiol and carboxylic acid group
(Acetyl CoA)
5- Guanidophosphates or phophagens is formed by the attachment of phosphate group
to guanidine group. (phosphocreatine)
ATP as universal energetic currency
ATP is adenosine triphosphate, contains nitrogen base adenine, pentose sugar and three
phosphate groups. The phosphate groups are linked by phosphoester and
phosphoalhydride bonds that produce high energy on hydrolysis
ATP is called as universal energy currency because the cell stores energy in the form of
ATP, it is the major molecule, and all the organisms and cells utilize the energy in both
catabolic and anabolic processes in the form of ATP
10. ATP synthesis: substrate-level phosphorylation and oxidative phosphorylation.
Examples of the substrate-level phosphorylation reactions.
ATP synthesis
Substrate level phosphorylation: transfer a phosphate group from a substrate directly
to ADP or GDP resulted production of ATP or GTP.
Glycolysis:
1,3 biphosphoglycerate + ADP ---phosphoglycerate kinase--> 3 phosphoglycerate + ATP
Phosphoenolpyruvate + ADP -----pyruvate kinase------> pyruvate + ATP
TCA cycle:
Succinyl CoA + ADP ------ succinyl CoA synthase-----à succinate + ATP
Oxidative phosphorylation: transporting electron chain to oxygen molecule to produce
H+ ion gradient which is essential for ATPase to synthesis of ATP in aerobic condition
11. Oxidative phosphorylation of ADP. Mechanism of coupling between oxidation and
phosphorylation. Mitchell’s chemiosmotic theory.
Oxidative phosphorylation of ADP: adding phosphate group from substrate to ADP and
form ATP
Mechanism between oxidation and phosphorylation: is well explained by
chemioosmotic hypothesis, the transfer of electrons through respiratory chain leads to
the pumping of protons from the matrix to the intermembrane space of mitochondria.
The proton concentration becomes higher on the cytosolic side and electrical potential
is generated there.
Mitchell’s chemiosmotic theory: is hypothesis stated that a proton-motive force was
responsible for driving the synthesis of ATP. During electron transport chain, H+ ion are
pumped to intermembrane space of mitochondria which leads to gradient different
between mitochondria matrix and intermembrane space.
pH gradient acts as battery for proton-motive force that is used to drive ATP synthesis.
12. Structure of electron-transport chain: components, complexes, arrangement in the
inner mitochondrial membrane. Structure of ATP-synthase. Mechanism of ATP
synthesis.
Structure of electron transport chain:
Components:
- NADH DH: is complex I
- Ubiquinone (coenzyme Q): lipid soluble and capable of diffuse within lipid bilayer of
mitochondria
- Succinate DH: is complex II
- Cytochrome c oxidoreductase is complex III
- Cytochrome c: soluble protein mediate transport between complex III and IV
- Cytochrome c oxidase: is complex IV
Complexes: 4 complex I, II, III, IV
- Complex I: entry point of electron from reduced NAD
- Complex II: entry point of electron from succinate
- Complex III: transfer electron from reduced cytochrome Q to cyochrome c (Complex
IV)
- Complex IV: carries electron to oxygen and reduces it to water
Arrangement
1 NADH – release 3H+ ion
1 FADH – release 2 H+
Structure of ATPase
ATP synthase is spherical projections on the matrix side of the inner mitochondrial
membrane, composed of 2 subunits:
- F0: is a proton pore that is embedded in the inner mitochondrial membrane and is
composed of four types of polypeptide subunits. It forms a channel or path through
which hydrogen ions may pass across the membrane. Hence, F0 is a proton channel of
inner mitochondrial membrane.
- F1: is bound to F0 sits on the matrix side of inner mitochondrial membrane. The F1 unit
is composed of 5 types of polypeptide subunits, and it contains the catalytic site for ATP
synthesis. It includes three beta-subunits, each of them has catalytic site for ATP
synthesis.
Mechanism of ATP synthesis
- F1 beta subunit has distinct opposite sites for ADP and Pi
- F0 acts as a channel for H+ ion to move down its concentration gradient
- Influx of H+ ion produce proton-motive force to rotate F1 and brings to
conformational change of beta subunit
- Beta subunit conformational change produce ATP from ADP and Pi
- Affinity of F1-ATP decreases. ATP is released to mitochondria matrix.
13. Regulation of oxidative phosphorylation. Uncoupling of oxidation and
phosphorylation. Physiologic implication. UCP-proteins. Molecular biological aspects of
mitochondrial function and dysfunction.
Regulation of oxidative phosphorylation:
1. Allosteric mechanism: level of ADP and free Pi in mitochondria matrix
2. Covalent modification of parathyroid hormone
3. Hormones: insulin, glucagon, epinephrine, norepinephrine
4. Calcium level
Besides, inhibitors of different levels of OP process:
- Complex I inhibitors: barbiturate drug (amytal), piericidine (antibiotic), rotenone
(insecticide)
- Complex III inhibitors: antimycin A and myxothiazol (antibiotic)
- Complex IV inhibitors: carbon monoxide, hydrogen sulfide, azide, cyanide
- ATPase inhibitors: oligomycin, cyanide, reotenone.
Uncoupling of oxidation and phosphorylation are certain chemical compounds that
allow normal function of electron transport chain without production of ATP
- Mechanism: disruption of H+ proton gradient by forming leakage channel of H+
proton. H+ not get to matrix through F0, no ATP is formed.
- Eg: dicumarol, FCCp, free fatty acids, 2,4 dinitrophenol
- Physiological implication: the dissipation of proton gradient is accompanied by
generating heat (brown adipose tissue - thermogenin) to maintain body temperature.
- UCP proteins: thermogenin in brown adipose tissue which contains a lot of
mitochondria helps generate heat after cold exposure.
Molecular biological aspects of mitochondria function and dysfunction:
- Mitochondria function can provide energy necessary for cell activities, sequester
calcium and detoxification of hydrogen peroxide, superoxide.
- Mitochondria dysfunction can lead to damage of DNA, proteins, membrane -> cell
death.
14. Reactive oxygen species and reactive nitrogen species. Nonezymatic and enzymatic
mechanisms of generation ROS and RNS. Lipid peroxidation: mechanisms, products.
Primary and secondary mechanisms of oxidative damage. Role of peroxidation in cell
death.
Reactive oxygen species: sharing 4 electron and form double bond between 2 O
molecules: O=O
Generation of ROS
Enzymatic (ETCh, Flavin – linked DH)
During process reduction of ubiquinone, arise semiquinone radical which is the most
important in generation other oxygen radical inside the cell. In respiratory arise so called
Flavin – linked DH take part again.
In NADPH oxidase reaction, flavin – linked DH is so important in the generation
superoxide anion radical. This reaction is very important with activated neutrophil.
Xanthine oxidase reaction: take part in such part hydrogen peroxide.
Reaction which catalyzed by oxidase of aa: flavin linked – DH take part in product of
reactive oxygen: hydroperoxide and it later take part in product of free radical reaction
inside the cell
2- Nonenzymatic:
Mechanism: coupled with outer oxidation of hemoglobin. Hemoglobin reduces iron in
the molecule of hemoglobin, use for reduction of oxygen. This oxygen may be
transferred with hemoglobin, and resulted from hemoglobin transform into
meghemoglobin and stimultaneously arise superoxide anion radical.
Fenton reaction Coupled with reduction of hydrogen peroxide . Hydrogen peroxide may
arise as product reduction of 2 electron of oxygen in the reaction catalyze by flavin
linked dehydrogenase. As source of additional electron, reduce iron arise hydroxyl
radical – the most free radical inside cell
Haber- Weiss reaction: this reaction coupled with reduction of hydrogen peroxide but
as reductant and use superoxide anion radical, resulted synthesis hydroxyl radical
Radiolysis of the water: water is elimination by X ray, Gamma ray, resulted hydroxyl
radical.
Reactive nitrogen species: nitric oxide derived compounds: peroxylnitrite, nitroxyl anion,
nitrosonium cation, etc
15. The antioxidant system of the body. Nonenzymatic antioxidants. Enzymatic
antioxidant defense system.
Antioxidants are defined as molecules which bring to decrease rate of free radical
processes
Antioxidant may be classified as
- Nonenzyme antioxidants: 3 groups
- Scavengers of free radical (vitamin C and E, ethanol, billrubine, melatonine,
carotinoids, etc)
- Chelators of transition valence metals (metalothionins, ferritin, transferrin, etc)
- Membrane stabilizer (vitamin E)
- Antioxidant enzymes (superoxide dismutase, catalase, peroxidases, etc)
Enzymatic antioxidant defence system: provide detoxification of free radical from toxic
to oxygen and water
16. Physiological importance of free radical oxidation. Free radical oxidation in
phagocytosis. Free radical oxidation in the pathology of cardiovascular system.
Physiological importance of free radical oxidation
Physiological role of oxidative stress is coupled with it participation in regulation
metabolic processes in adaption organism to effect of adverse factors. It takes part in:
1- Signal transduction
2- Gene expression (regulation expression of stress protein)
3- Synthesis of biological active substances – prostaglandins, leukotrienes and
cytokinase.
4- Renovation of membranes and intracellular proteins
5- Antimicrobial defense of organism
Resulted from those effects oxidative stress take part in regulation inflammation,
growth, proliferation, apoptosis, and antimicrobial defense.
Free radical oxidation in phagocytosis
- Both ROS and RNS can damage nucleic acids (DNA and RNA), membrane lipids,
proteins which are cells main components.
Free radical oxidation in the pathology of cardiovascular system
1- Appear tissue hypoxia – increase free radical production by mitochondrial respiratory
chain
2- Activation of xanthine oxidase
3- Activation of NADPH-oxidase
4- Increase production of epinephrine
5- Inhibition of antioxidant enzymes
MODULE III. METABOLISM OF CARBOHYDRATES
1. The biological role of carbohydrates. The daily requirement for carbohydrates in
adults and in children. Dietary carbohydrates of animal and vegetable origin.
Biological role of carbohydrates:
- Energetic: carbohydrate breakage is accompanied by release energy which can be used
by cells for various purposes
- Structural: carbohydrates take part in the creating intracellular matrix
- Carbohydrates take part in the defence of human body from infection agent. (structure
of antibodies)
- Carbohydrates are in the structure of different cell receptor which function in
regulation thus take part in cell signaling and cell-cell recognition processes.
Daily requirement of carbohydrates
- In children: 100g per day
- In adult: 200-300g per day
Carbohydrates sources
- Animal origin: milk, cheese, dairy products
- Vegetable origin: cereal, rice, corn, potato, sugar cane
2. Structure and function of carbohydrates: monosaccharides, disaccharides, oligo and
polisaccharides. Derivatives of monosaccharides - acetylhexosamine, glucuronic acid.
Monosaccharides: contain single residue of polyhydroxy carbonyl. The formula of
structure is Cm(H2o)n.
Types of monosaccharides: fructose, glucose, mannose, ribose, galactose
Derivatives of monosaccharides:
- Acetyl hexosamine: derivative of glucose, consists of glucosamine and acetic acid.
- Glucuronic acid: derivative of glucose, with 6th carbon oxidized to carboxylic acid. It is
important in detoxification in human body
Disaccharides: formed from 2 monosaccharides linked by glycosidic bond. Types:
sucrose, lactose, maltose
Oligosaccharide: consists of 2-10 monosaccharides linked by glycosidic bond.
Polysaccharide: are linear or branched polymers of monosaccharides and its derivatives.
2 groups
- Homopolysaccharides: contain identical monosaccharides as monomers: glycogen,
starch, dextran, cellulose
- Heteropolysaccharides: contain different monosaccharides and their derivatives as
monomers: glycosaminoglycan (hyaluronic acid, dermatan sulfate)
3. Digestion of carbohydrates. Enzymes in carbohydrate digestion: salivary α-amylase,
pancreatic α-amylase, enzyme complexes in small intestine.
Digestion of carbohydrates: the breakdown of carbohydrates to monomers for
absorption first start in oral cavity then stomach and absorption in intestine.
A-amylase: brake carbohydrates into short-branched oligosaccharides (pH optimal = 7.5)
- Salivary a-amylase: is the amylase that is produced by the salivary glands and works in
mouth, act on raw form of carbohydrates (from starch to di and monosaccharides)
- Pancreatic a-amylase: is the amylase that is produced by acinar cells in pancreas, works
in stomach and small intestine, act on complex carbohydrates that enter into the
stomach after partial digestion.
Enzyme complexes in small intestine: product: oligosaccharides
4. Malabsorption of carbohydrates: biochemical mechanisms. Sucrose and lactose
intolerance: biochemical causes, consequences and mechanisms of development of
symptoms.
Malabsorption syndrome refers to several disorders in which the small intestine can’t
absorb enough of certain nutrients and fluids: macronutrients (proteins, carbohydrates,
and fats), micronutrients (vitamins and minerals), or both.
Biochemical mechanisms:
- Enzyme deficiencies: stomach is not able to form digestive enzymes to break down food
- Disorder in enterocytes: nutrients cant get absorbed due to error in channels, etc...
Different causes: - Infective agents, diseases, parasites, … or Inflammation of mucosa,
disorder synthesis transporters
Lactose intolerance: decreases ability of digestion of dairy product due to the lack of
enzyme lactase in small intestine to break down lactose to glucose and galactose.
Lactose: (milk sugar) disaccharide cannot be absorbed directly through wall of small
intestine into blood stream, so in absence of lactase, food pass through colon and bacteria
in colon can metabolize lactose, resulting fermentation, produces amounts of H2, CO2,
methane)
Symptoms: abdominal discomfort and diarrhea
Lactose: broken by lactase into galactose and glucose
Sucrose intolerance: sucrase-isomaltase enzyme cannot be produced to catalyze
metabolism reaction of sucrose and starch. (caused: genetic mutations, explain why some
people are better able to tolerate starch in diet than others)
Sucrose: broken by sucrose into glucose and fructose, has high sweet taste, obligate
compound of food, its oligosaccharide.
Symptoms:
Diagnosis:
- Fecal analysis: lactose and sucrose can’t be digested pass out with feces.
- Breath test: lactose malabsorption caused a high level of H2 in breath.
- Blood test: after eating, the glucose level in blood does not increase.
5. Transport of monosaccharides through membranes: facilitated diffusion and active
transport. Glucose transporters: types, structures, functions. Insulin-dependent glucose
transporters.
Transport of monosaccharides through membrane:
- Active transport: ensure that glucose flows to intestinal cells and blood stream no
matter how the concentration of glucose is.
- Facilitated diffusion: (mainly)
+ After eating, the concentration of glucose in gut will be higher than in the blood,
glucose flow downhill gradient to the bloodstream.
+ When then intestine is empty, glucose concentration in it is lower than in the
blood, glucose flow downhill from bloodstream to the gut.
Glucose transporter is a special protein, with many types of cells in different tissues,
organs (epithelium, liver cell, fat cell, …). 2 types:
- Glucose dependent: in muscle, fat cell
- Glucose independent: not sensitive to insulin
Glucose
Localization
Structure
Function
transporter
RBC, brain, kidney, colon, retina,
Consist of 1 long
GLUT – 1
Uptake glucose
placenta
polypeptide chain,
GLUT – 2
GLUT – 3
which contain many
Rapid uptake and Serosal surface of intestinal cells,
a-helix parts. A helix
liver, b-cell of pancreas
release of glucose
part can contact
Neuron, brain
hydrophobic part, Uptake of glucose
GLUT – 4
GLUT – 5
SGLT – 1
which helps binding
glucose to binding
site and change the
conformation of the
protein and release
the
glucose
to
extracellular space.
Insulin stimulated Skeletal and heart muscle, adipose
tissue
uptake glucose
Absorption
of Small intestine, testis, sperms, kidney
glucose
Active uptake and
reabsorbtion
of Intestine, kidney
glucose
Insulin dependent transporters: GLUT4. The insulin-regulated glucose transporter found
primarily in adipose tissues and striated muscle (skeletal and cardiac).
At the cell surface, GLUT4 permits the facilitated diffusion of circulating glucose down its
concentration gradient into muscle and fat cells
Localization: skeletal, heart muscle, adipose tissue
6. Glucose pathways. Glucose phosphorylation. The role of glucose-6-phosphate.
Glucose pathways:
Metabolic pathways of glucose include multiple processes: glycolysis, gluconeogenesis,
glycogenolysis and glycogenesis, pentose pathway
Catabolism of glucose: contains 2 processes:
- Glycolysis: converts 1 glucose into 2 molecules pyruvate
- Glycogenolysis: breaks down glycogen into glucose
Glucose phosphorylation: an important irreversible reaction in metabolism of glucose. It
transforms glucose into glucose 6 phosphate under catalyse of hexokinase (in muscle)
or glucose kinase (in the liver). This reaction expends 1 ATP
Glucose 6 phosphate is the first intermediate of glucose metabolism, it plays central
role in connect glycolysis, glycogenesis, pentose phosphate pathway, lipogenesis.
- G6P cannot move out of the cell. Forming of G6P prevent the release of glucose into
bloodstream.
7. Glucose metabolism in liver: role of glucokinase and glucose-6 phosphatase in the
maintenance of a constant concentration of glucose in the blood.
Glucose metabolism in liver: glycolysis and gluconeogenesis.
Phosphorylation and dephosphorylation of glucose are irreversible reactions.
- Glycolysis: glucokinase phosphorylate glucose into G6P which is first step of
breakdown glucose to form pyruvate.
- Gluconeogenesis: G6Pase catalyses the last reaction forming glucose from noncarbohydrate substrates.
Glucose can pass through cell membrane but G6P cannot. Glucose kinase and G6Pase
under influence of insulin and glucagon can maintain the constant level of glucose in
the blood by uptake or secrete glucose from the cell.
8. Glycogen synthesis from glucose-6-phosphate. Biologic role, reactions, enzymes.
Tissue and cellular localization.
Glycogenesis:
Glycogen synthesis located in cytoplasm of muscle tissue, hepatocytes and adipose
tissues for storage glucose under hyperglycemia condition
- Phosphoglucomutase transfer phosphate group from
position 6th carbon to 1st carbon, converting G6P to G1P
UTP (uridine triphosphate) composes of 3 Pi.
- Under catalysation of pyrophosphorylase, UTP releases
2 Pi and bind to G1P forming UDP glucose (UDPG)
Pyrophosphate = 2 Pi = take away 2 Pi from UTP
- UDPG interacts with glycogenin (auto glycosylation).
- Glycogen synthase keeps adding UDPG to the chain,
forming 1,4 glycogen chain.
- Branching enzyme transfer the linkage between 1-4th
carbon to 1-6th for further branching of glycogen.
9. Degradation of glycogen. Biologic role, reactions, enzymes. Tissue and cellular
localization.
Glycolysis takes place in cytoplasm of muscle, liver, or adipose tissues, forming and
releasing glucose to maintain normal blood glucose level in hypoglycemia condition
- Glycogen phosphorylase catalyzes the phosphorylation of 1,4 glycosidic bond of
glycogen, releasing free glucose 1 phosphate
- Debranching enzyme breakdown 1,6 glycosidic bond and release free glucose
10. Features of glycogen metabolism in liver and muscle under certain physiological
conditions (food intake, fasting, muscle activity). Hormonal regulation of these
processes.
- During food intake process, glucose level in the blood increases (hyperglycemia). Liver
and muscle take up glucose (to maintain normal blood glucose level) and synthesis
glycogen (glycogenesis). This process is influence by action of INSULIN
- During fasting or muscle activity, glucose level in the blood decreases (hypoglycemia).
Liver and muscle break down glycogen (glycogenolysis) and release free glucose to the
blood to bring back normal level of the blood. This process is influence by action of
GLUCAGON, EPINEPHRINE, NOREPHINEPHRINE
11. Regulation of enzyme activity in metabolism of glycogen - glycogen synthase and
glycogen phosphorylase: hormonal regulation - the effect of epinephrine and glucagon
(adenylate cyclase mechanism, the role of cyclic AMP and protein kinase A); the role of
insulin and phosphodiesterase in decreasing of cAMP concentration in the cell; allosteric
regulation of glycogen phosphorylase activity with the participation of AMP; calciumdependent activation of glycogen phosphorylase kinase.
Glycogen synthase (glycogenesis) and glycogen phosphorylase (glycogenolysis) catalyze
for reverse process of glucose and glycogen.
Hormonal regulation:
Epinephrine and glucagon influence glycogenolysis to increase glucose blood level.
Insulin and phosphodiesterase: decrease cAMP level
Allosteric regulation of glycogen metabolism:
- ­ATP -> ¯AMP. ­AMP stimulate glycogen phosphorylase
- - Ca2+ activates phosphorylase kinase which is activator for transform of glycogen
phosphorylase from inactive to active state
12. Genetic disorders of glycogen metabolism: glycogen synthesis and glycogen
degradation (liver, muscle and mixed glycogenoses).
Genetic disorders that lead to deficiency of glycogen metabolism enzymes: GLYCOGEN
STORAGE DISEASE
- Hers disease: defective in liver glycogen phosphorylase deficiency
- Glycogen storage disease type VIII and IX: deficiency of phosphorylase kinase b
- Pompe disease: deficiency of lysosomal acid alpha 1,4 glucosidae
- Cori disease: deficiency of glycogen debranching enzyme (alpha 1,6 glycosidic bond)
- Andersen disease: deficiency of glycogen branching enzyme
- McArdle disease: deficiency of muscle glycogen phosphorylase
13. Metabolic pathways of glucose. The role of glucose-6-phosphate.
- Under phosphorylation, glucose in transform into G6P which plays central role in
carbohydrate metabolism:
+ Glycogenolysis, glycogenesis
+ Pentose phosphate pathway
+ Glycolysis, gluconeogenesis -> citric acid cycle -> electron transport chain to produce
ATP
14. The process of glycolysis: localization and conditions, the sequence of reactions and
enzymes, the final products, involvement of adenine nucleotides and energy effect,
irreversible reactions of glycolysis, reactions associated with consumption of ATP,
substrate-level phosphorylation reactions, their role, glycolytic oxidoreduction.
Glycolysis: takes place in cytoplasm of cells to produce energy for living activities
- Glucose gets into cell through GLUT
Final products of glycolysis: 2 pyruvate 2 NADH 4 ATP
Substrate level phosphorylation: substrate of glycolysis donates a phosphate to ADP to
produce ATP (last reaction: PEP -> pyruvate)
Glycolytic oxidoreduction reaction: oxidoreduction of NAD+ and glycerol 3 phosphate
to produce NADH essential for electron transport chain to produce ATP (glyceraldehyde
3 phosphate -> 1,3 biphosphate)
15. The process of gluconeogenesis: localization and conditions of the reactions, the
substrates, the sequence of reactions and enzymes, reactions associated with the
consumption of GTP and ATP, irreversible reactions of gluconeogenesis, the role of
gluconeogenesis in fasting and in physical exercises, energy consumption for the
synthesis of one molecule of glucose.
Gluconeogenesis occurs in liver or kidney (PCT) to provide energy mainly for the brain
during fasting (hypoglycemia)
Substrates: lactic acid, glycerol, amino acids, fatty acids
- Lactic acid (product of muscle, released to the bloodstream and get to the liver) is
converted to pyruvate by lactate dehydrogenase
- Glycerol and fatty acids are products of TAG hydrolysis.
+ Glycerol - glycerol phosphatase-> glycerol 3 phosphate (G3P) (in cytoplasm)
+ G3P shuttle (in mitochondria): G3P -glycerol 3 phosphate DH -> dihydroxyl acetone
phosphate, coupled with generate NAD+
- Amino acids: amino acid under transamination/deamination, forming keto acid that
can go to TCA and join gluconeogenesis
Reactions associated with consumption of GTP and ATP: total 6 ATP
- Phosphorylation of glycerol 3-phosphate to glycerol 1,3 phosphate
2 ATP
- Carboxylation of pyruvate forming oxaloacetate
2 ATP
- Conversion of oxaloacetate to phosphoenolpyruvate
2 ATP
Irreversible reactions: bypass I, II, III
- Pyruvate (-> oxaloacetate) -> phosphoenolpyruvate (PEP)
Pyruvate cannot convert to PEP directly, so it has to be converted to oxaloacetate (in
TCA) then oxaloacetate -phosphoenolpyruvate carboxyl kinase (PEPCK)-> PEP.
+ MALATE-ASPARTATE SHUTTLE: Pyruvate is converted to OAA through TCA in
mitochondria. OAA can’t pass through mitochondria to cytoplasm so it is
transformed to malate. After getting to cytoplasm, malate -> OAA
- Fructose 1,6 phosphate -fructose 1,6 biphosphatase-> Fructose 6 phosphate
- G6P - G6Pase (smooth endoplasmic reticulum)-> glucose
During fasting and in physical exercises, blood glucose level decrease. Gluconeogenesis
generate glucose necessary for ATP synthesis mainly for the brain (highly required ATP)
and maintain glucose level in the blood when sufficient carbohydrate is not available
from diet or glycogen is reserves
Gluconeogensis is a mechanism to clear the products of metabolism from other tissues:
lactate, glycerol, etc
16. The reciprocal regulation of glycolysis and gluconeogenesis: hormonal regulation the role of insulin, epinephrine, cortisol, glucagon; allosteric regulation - role of ATP,
ADP, AMP, citrate, fatty acids, glucose-6-phosphate, fructose-6-phosphate, fructose-1,6diphosphate, acetyl SCoA.
Glycolysis and gluconeogenesis are opposite process.
Hormonal regulation:
Glycolysis
Insulin
(-)
Glucagon
(+)
Epinephrine (+)
Cortisol
(-)
Allosteric regulation:
Gluconeogenesis
(+)
Insulin
(-)
Glucagon
(-)
Epinephrine
(+)
Cortisol Stimulate synthesis of PEP carboxykinase
17. Anaerobic oxidation of glucose. Pathways for glycolysis products under anaerobic
conditions. Energy effect of the oxidation of glucose and glycogen under anaerobic
conditions.
Anaerobic oxidation of glucose is the process forming lactate in the limited amount of
O2.
Pathway for glycolysis under anaerobic conditions:
Glycogen -> G1P -> G6P -> Pyruvate -> Lactate (then transferred to the liver)
Under anaerobic conditions, glycolysis forms 2 ATP and utilizes 6 ATP to generate
glucose molecule.
18. Pathways for products of glycolysis under aerobic conditions. Glycerol 3-phosphate
shuttle and malate–aspartate shuttle. Energetic effect of aerobic glucose oxidation.
Products of glycolysis: 2 pyruvate. 2 pathways
- Anaerobic pathway: -lactic DH-> lactic acid. Lactic acid can decrease blood pH and be
transferred to liver for gluconeogenesis.
- Aerobic pathway: -pyruvate decarboxylase-> acetyl CoA then goes to TCA, oxidative
phosphorylation to create ATP
Glycerol 3 phosphate shuttle: Necessary for utilize glycerol for gluconeogenesis
Glycerol – glycerol phosphatase-> glycerol 3 phosphate (G3P) (in cytoplasm)
G3P <-glycerol 3 phosphate DH -> dihydroxyl acetone phosphate (in mitochondria)
- DHAP accepts electron from NADH to become a-glycerol phosphate. A-glycerol
phosphate enters mitochondria where it is oxidized to form FADH2 and DHAP.
- This shuttle produces 30 ATP per molecule of glucose metabolized
Malate-aspartate shuttle: important for gluconeogenesis (reverse of bypass I: pyruvate > PEP)
- Oxaloacetate accepts electron from NADH to become malate. Malate enters
mitochondria and oxidizes to OAA and NADH. This shuttle produces 32 ATP per
molecule of glucose metabolized.
Aerobic glucose oxidation can provide 36 ATP (in total) per 1 molecule of glucose,
compare to anaerobic pathway (2ATP)
19. Stages of aerobic oxidation of glucose, and the summary reaction of aerobic
breakdown of glucose. Advantages of aerobic oxidation of glucose.
Aerobic oxidation of glucose is the process breaking down of glucose forming energy.
There are 4 stages:
-
Glycolysis: 1 glucose 2 pyruvate
Pyruvate oxidation: 2 pyruvate 2 acetyl CoA
Citric acid cycle: produces 1 ATP, 3 NADH, 1 FADH2, releases CO2
Oxidative phosphorylation: from NADH and FADH2, taking in O2 and making
ATP and produces water
C6H12O6 + O2 → 6CO2 + 6H2O + 2870kJ
Advantages: most efficient way for utilize glucose
20. Pyruvate: metabolic pathways, biologic role, reaction of conversion to acetyl-ScoA
and oxaloacetate, the energy balance of oxidation to CO2 and H2O.
Pyruvate occupies an important junction in the carbohydrate catabolism
Glucose
↑
Alanine
↔
Pyruvate
↓
Acetyl CoA
↔ Lactate
↓
Malate
Transition: conversion of pyruvate to acetyl CoA under catalysed of pyruvate
decarboxylase
TCA cycle: acetyl CoA combines with oxaloacetate then forming last product is OAA
During oxidation of glucose, TCA cycle forming 4 CO2 and 1 ATP, and electron transport
chain forming H2O and 38 ATP
21. Vitamins as coenzymes in pyruvate metabolism (H, В1, В2, В3, В5): dietary sources,
daily requirements, biochemical functions, signs of deficiency.
VItamin
Chemical
structure
Thiamin
(B1)
Thiamin
Riboflavi
n (B2)
Pantothe
nic acid
(B5)
Niacin
(B3)
Biotin
Riboflavin
Daily
requirement
(for adult)
1.0-1.5
mg/day
1.2-1,7
mg/day
B-alanine
and
pantoic
acid
Nicotinic
13-19
acid and
mg/day
nicotinami
de
Biotin
Coenzy
me
form
Thiamin
pyroph
osphate
(TPP)
(by
thiamin
diphosp
hate
kinase)
Precurs
or for
FMN
and
FAD
Precurs
or of
NAD+
and
NADP+
Significance
Manifestation
of vitamin
deficiency
B1 reacts with ATP to form TPP which Weight loss,
is necessary for
appetite
-pyruvate dehydrogenase
suppression,
– a.ketoglutarate dehydrogenase
nausea, mental
catalyzed reactions in carbohydrate
depression,
metabolism
peripheral
-transketolase, an enzyme in the
neuropathy,
pentose phosphate pathway
fatigue.
Help convert carbohydrates, fats,
protein into energy
Take part in wide range of redox
reaction:
• Succinate dehydrogenase
• Xanthine oxidase
Help convert carbohydrates, fats,
protein into energy
Is required for synthesis of coA
(component of acyl carrier protein)
Help convert carbohydrates, fats,
protein into energy
Is required to synthesis of active
forms of vitamin B3 – NAD+, NADP+
(funcions: cofactor for
dehydrogenase)
Not a true vitamin (because NAD+
can be derived from aa tryptophan.
Cofactor required for enzyme
involved in carboxylation reactions
(acetyl CoA carboxylase and pyruvate
carboxylase
Angular
stomatitis and
cheilosis,
bloodshot
seborrhea,
trembling,
sluggishness,
photophobia
Resemble to
those other vit
B deficiencies.
Glossitis,
dermatitis,
weight loss,
diahrrea,
depression
and dementia
Muscle pain,
loss of
appetite,
depression,
grayish skin
color
22. Glucose-lactate cycle (Cori cycle). Biologic role in physical exercises. Sources of
lactate in human organism.
Cori cycle
- Muscle has no G6Pase to convert G6P to glucose to utilize, so it is converted to
pyruvate.
- During muscle work, anaerobic conditions (lacks oxygen) can convert pyruvate
(product of glycogenolysis) to lactate.
- Lactate cannot generate energy for muscle work in the muscle so it is released to the
bloodstream to the liver for gluconeogenesis
- ROLE: allow muscle to function anaerobically by removing excess substances
23 Glucose-alanine cycle. Biologic role in physical exercises and in fasting.
- Muscle has no G6Pase to convert G6P to glucose to utilize, so it is converted to
pyruvate.
- In aerobic condition transamination of pyruvate and glutamate forming alanine.
- Alanine can’t be utilized in the muscle, it travels from skeletal through the blood to the
liver
- In the liver, alanine combines with alpha ketoglutarate to form pyruvate and
glutamate. Pyruvate is essential for gluconeogenesis and glutamate under oxidative
deamination release ammonia to urea cycle
- During physical exercise and fasting, this cycle takes up amino acids from the skeletal
muscle to the liver for metabolism to provide energy and maintain blood glucose.
24. The effect of ethanol on metabolism of carbohydrates in the human body. Causes of
hyperlactatemia and hypoglycemia in alcohol intoxication.
Effects of ethanol on carbohydrate metabolism
Ethanol inhibits insulin expression that leads to hyperglycemia
Ethanol inhibits both gluconeogenesis and glycolysis that leads to hypoglycemia
Besides, ethanol inhibits galactose metabolism by inhibiting the key enzyme uridine
diphosphate galactose 4-epimerase. Ethanol inhibits the metabolism of fructose and
sorbitol.
Hyperlactatemia and hypoglycemia: the decreasing of glucose and increasing of lactate
in the blood due to alcohol intoxification.
- Consumption and metabolism of ethanol -> high levels of NADH in the cytosol and
mitochondria of the liver cell. High levels of NADH stimulate the synthesis of lactate
from pyruvate and decrease glucose level -> hypoglycemia and hyperlactatemia
25. Features of glucose metabolism in the liver, brain, skeletal muscle, adipose tissue,
erythrocytes.
- Liver: main site for all glucose metabolism, regulates blood glucose level
- Brain: glucose is main source for brain. Within the brain, glucose is either oxidized to
ATP or synthesize glycogen.
- Skeletal muscle: glucose is used in anaerobic or aerobic pathway to create energy or
glycogen synthesis
- Adipose tissue: uptake glucose to form glycerol 3-phosphate (glycolysis), which is
essential for synthesis of TAG.
- Erythrocytes: lacks mitochondria, after glycolysis, pyruvate is formed and converted
into lactate (no O2 condition) or forming 2,3 biphosphoglycerate. Besides, glucose gets
to pentose phosphate pathway forming ribose 5 phosphate
26. Characteristics of the pentose phosphate pathway of glucose oxidation: localization
and role of the pentose phosphate pathway, reactions of the oxidative phase, principles
of nonoxidative phase, enzymes, coenzymes, interaction with glycolysis, the role of the
pentose phosphate pathway in adipose cells, in erythrocytes, in dividing cells.
PPP takes place in liver, mammal glands, adipose tissues, adrenal cortex and rapid
dividing cells. Enzymes of this pathway are located in cytoplasm of cells
Products of PPP: NADPH, ribose 5 phosphate and glycolytic intermediates. Depend on
demand and condition of the body, PPP and glycolysis can supply sufficient glucose,
ATP, NADPH, or ribose 5 phosphate (for DNA, RNA synthesis, etc)
OXIDATIVE PHASE: forming NADPH and ribose 5 phosphate
NON OXIDATIVE PHASE
Principles:
- Using ribose 5 phosphate forming glycolytic substrates: GA3P and F6P for glycolysis
- Not using, not forming ATP
Role of PPP in different tissues:
RBC: production of reduced NADP (antioxidant activity)
Hepatocytes: production of reduced NADP (antioxidant activity, detoxification, anabolic
processes); production of ribose-5-phosphate (anabolic processes)
Adipocytes: production of reduced NADP (lipogenesis)
Mammal gland: production of reduced NADP (lipogenesis)
Adrenal cortex: production of reduced NADP (steroidogenesis)
Fast division cells: production of reduced NADP and ribose 5-phosphate
(membranogenesis, synthesis of nucleic acids)
27. Hereditary deficiency of glucose-6-phosphate dehydrogenase. The factors that
trigger manifestations of the disease.
G6PD catalyzes first reaction of PPP. This pathway helps red blood cells work (by
produce NADPH for antioxidant) and synthesis of DNA, RNA (ribose 5P).
Deficiency of G6DP leads to anemia hemolysis (destruction of erythrocytes) and it can
leads to jaundice, gallstones.
This is caused by genetic disorder or reactions of body to certain drugs (aspirin high
dose, quinidine, antibiotics like quinolones, nitrofurantoin), stress, infection.
28. The conversion of fructose into glucose. Pathways of fructose metabolism. Disorders
of fructose metabolism. Differences in metabolism of fructose in the liver and in
muscles. Polyol pathway of fructose synthesis pathway, biologic role.
Conversion of fructose into glucose: in hepatocyte, where have both fructokinase and
glucose 6 phosphatase
Fructose metabolism: take place in hepatocyte
- Fructose which is product of sucrose breaking down and absorption through intestine
is converted to intermediate of glycolysis
- Fructose metabolism forming pyruvate even faster than glucose
Disorders in fructose metabolism:
Fructosuria: disorder caused by deficiency of fructokinase. This disorder decreases
ability to utilize fructose of the body
- Manifestation: normally asymptomatic.
- Treatment: no need
Fructose intolerance: disorder caused by deficiency of fructose 1 phosphate aldolase b
- Deficiency of this enzyme rapidly leads to accumulation of F1P which is toxic to the
liver, thereby leading to impaired glycolysis, glycogenolysis and gluconeogenesis.
- Manifestation: asymptomatic until digestion of fructose or sucrose food. Jaundice,
vomiting.
- Complications: cirrhosis, kidney failure
- Treatment: elimination of all sources of sucrose, fructose, and sorbitol.
Differences of fructose metabolism in liver and muscle
- In the liver, enzyme fructokinase catalyzes conversion of fructose to fructose 1
phosphate. Then F1P can be converted to glycolytic intermediate forming pyruvate or
glucose
- In the muscle, there is no fructokinase. Hexokinase stimulates conversion of fructose
into F6P which is glycolytic intermediate to produce energy for muscle work.
Polyol pathway of fructose synthesis:
- G->F: take place in liver, ovary, and seminal vesicle cells
- G->S: take place in Schwann cells, retina and kidney cells where lacks sorbitol DH
- In lack of hexokinase condition, this process can take up excess glucose forming
fructose or sorbitol to re-enter to glycolysis pathway. These reactions don’t require ATP.
29. Role of galactose in the body. Galactose metabolism. Key enzymes of galactose
metabolism. Galactosemia, molecular causes, clinical manifestations and principles of
treatment.
Galactose is product from breaking down of lactose (in dairy products)
Galactose main pathways:
- Transformation into glucose (in liver, kidney cortex, contains G6Pase → glucose)
- Glycolysis (energy supplement of the cell) (galactose → glu6P)
- Biosynthesis of micropolysaccharides (galactose amin, acetyl galactoseamin →
galactorunic acid) forming main compound of extracellular matrix.
Galactose metabolism:
Key enzymes of galactose metabolism: important for changing galactose to glucose
residue
- Galactokinase (1st reaction)
- Galactose 1 phosphate Uridyl transferase (2nd reaction)
Deficiency of these enzyme can lead to cataract, serious influence to organs
Galactosemia: disorder of deficiency of galactose 1 phosphate uridyl transferase
- Manifestation: cataract, jaundice, mental retarded, feeding difficulties, poor gain
weight
- Principles of treatment: elimination galactose from diet
30. Glucose metabolism in erythrocyte: glycolysis, pentose phosphate pathway, 2,3
bisphosphoglycerate synthesis.
There are 2 pathways of glucose metabolism in erythrocyte:
Glycolysis:
- Main pathway in RBCs. Since RBCs lack mitochondria, the end product is lactic acid.
This process gains 2 ATP help RBC maintain shape and flexibility.
- Glycolysis forms 1,3 biphospho-glycerate (1,3 BPG) then 2,3 BPG which binds to Hb
decreasing affinity for O2 (regulator of Hb-O2 affinity)
- PPP: provides NADPH to keep glutathione in reduced state which has important role in
destruction of peroxide. Peroxide cause irreversible damage to membranes, DNA and
must be removed to prevent cell damage and death
2,3BPG synthesis:
31. Hormonal regulation of carbohydrate metabolism. Influence of insulin, glucagon,
epinephrine, cortisol on blood glucose level and on intracellular glucose metabolism.
Insulin- dependent tissues. Hormone-sensitive enzymes of carbohydrate metabolism,
mechanisms of their regulation.
Blood glucose level
Insulin
¯
Glucagon
­
Epinephrine
­
Glucose metabolism
Glycogenesis, PPP, glycolysis
Glycogenolysis, gluconeogenesis
Glycogenolysis, glyconeogenesis
­
Cortisol
Gluconeogenesis (­PEP carboxyl kinase)
Insulin-dependent tissues: liver, muscles, adipose tissue,
Hormone-sensitive enzymes: key enzyme of carbohydrate metabolism
- Eg: glycogen synthase, glycogen phosphorylase, phosphofructokinase, fructose
biphosphotase, glucose kinase, pyruvate kinase, etc
- Insulin/glucagon activates 2nd messenger (insulin: PIP3 -> tyrosine protein kinase;
glucagon: cAMP -> protein kinase A) that can phosphorylation/dephosphorylation for
enzyme activation
32. The physiological and pathological hyper- and hypoglycemia.
Physiologic
Pathologic
Hyperglycemia
Resulted from decreasing insulin,
decreasing ability to store glucose
of the body
-
Mild hyperglycemia: no
symptoms
Severe hyperglycemia: increase
urine output, fatigue, muscle
weakness, low blood pressure,
impaired CNS
Hypoglycemia
Resulted from meal skipping or
overdose of insulin leads to high
storage of glucose and low
glucose blood level
- Mild hypoglycemia: hunger,
rapid pulse, anxiety
(sympathoadrenal symptoms)
- Severe hypoglycemia: seizure,
blur vision, faint, coma
33. General characteristics of diabetes type 1 and 2. Disorders of carbohydrate
metabolism. Biochemical mechanisms of diabetes complications.
Type I diabetes mellitus: endocrine disease by disordered of carbohydrate metabolism:
inability of pancreas to produce insulin
Results:
- Glucose uptake by tissues is impaired, despite the fact that glucose is being released
by the liver
- Glycolysis is depressed
- Gluconeogenesis is stimulated
- Glycogen storage is depleted
Manifestations:
- Hyperglycemia (high level of glucose in blood)
- Glucosuria (excretion of glucose with urine)
- Increased urinary output
Treatment: addition of insulin to maintain normal glucose blood level
Type II diabetes mellitus: insulin is unable to interact with glucose (insulin resistance)
Results: the same
Manifestation: the same
Treatment: insulin injections, use medication for better insulin working.
MODULE IV. METABOLISM OF LIPIDS
1. Classification of lipids. Polyunsaturated fatty acids: w-6 (linoleic, g-linolenic,
arachidonic acid), w-3 (a-linolenic). Chemical structure. Biologic role. Vitamin F.
Triacylglycerols, structure, biological role and function. Structure of phospholipids:
phosphatidylserin, phosphatidylcholine, phosphatidylinositol,
phosphatidylethanolamine. Biologic role.
Classification of lipids:
- Simple lipids: highly hydrophobic, don’t contain hydrophilic part:
+ TAG and their derivatives
+ Waxes
+ Sterols
+ Terpens
- Complex (conjugated) lipids contain hydrophilic component in their structure:
+ Phospholipids
+ Glycolipids
+ Lipoproteins
Polyunsaturated fatty acids: essential fatty acids from plants
(w-6) linoleic C17H31COOH contain 2 double bonds
((w-6) g-linolenic: C17H29COOH contain 3 double bonds
(w-6) arachidonic acid: C19H31COOH contain 4 double bonds
(w-3) a-linolenic: C17H29COOH contain 3 double bonds
Chemical structure: hydrophobic nature. Fatty acids are water insoluble long-chain
hydrocarbons with one carboxyl group at the end of the chain. Fatty acids can be
saturated or unsaturated.
Biological role:
- Storage form of metabolic fuel
- Transport form of metabolic fuel
- Structural component of biological membranes
- Regulator of metabolism
- Defence of organism against mechanical impacts
- Take part in the thermoregulation
Vitamin F:
- Complex of polygenic fatty acids (polyunsaturated fatty acids): alpha linolenic acid
(ALA) and linoleic acid (LA
- Sources: plant oil, fish, milk, dairy product
- Significance: membrane phospholipid (cell membrane component); increase
polyunsaturated fatty acid -> increase properties of cell membrane; precursor of some
active substances in human body (bile)
- Hypoviaminosis: disorder genetic factor, reproduction factor, atherosclerosis
Triacylglycerol: nonpolar, hydrophobic and insoluble
Structure: 1 glycerol and 3 fatty acids linked by ester bonds
Biological role:
- Storage of energy in adipose tissue
- Lipid homeostasis and growth
Function:
- Energy storage and providing
Phospholipids: a lipid made of glycerol, 2 fatty acids and phosphate linked head group.
Phosphatidylserin: contain glycerol backbone, 2 fatty acid tails and a phosphate group
substituted with and serine.
- Play key role in cell cycle signalling, component of cell membrane
- Increasing thrombin level
Phosphatidylcholin: contain glycerol backbone, 2 fatty acid tails and a phosphate group
substituted with choline
- Major constituent of cell membranes and pulmonary surfactant
Phosphatidylinositol: contain glycerol backbone, 2 fatty acid tails and a phosphate group
substituted with an inositol polar head group
- Play role in lipid signalling, cell signalling and membrane trafficking
Phosphatidylethanolamine: contain glycerol backbone, 2 fatty acid tails and a phosphate
group substituted with ethanol amine group
- Forming white mater in nervous system
- Play role in membrane fusion
- Increasing thrombin level
2. Digestion of lipids. Dietary sources of lipids, daily consumption of lipids. The stages of
digestion of lipids in gastro-intestinal tract.
Digestion of lipids:
- Begin in duodenum. Lipids undergo effect of pancreatic hydrolytic enzyme. Pancreatic
lipase is secreted into duodenum in inactive form. They are activated by bile acids. Bile
acids help lipid form emulsification for enzymatic hydrolysis.
Dietary sources: from animal oil and vegetable oil
Daily requirement: 40-70g per day
Stages of lipid in gastro-intestinal tract:
- In stomach: breaking down of lipid into small pieces
- In duodenum: emulsification of lipid by bile acids and mixing with enzyme.
- In intestine: absorption of fatty acids and glycerol in epithelial cells and being
resynthesis
- In enterocyte: forming lipoprotein for transport of lipid in the bloodstream.
3. The composition of bile and its role. Types of bile acids, their functions, structure.
Disorders of bile secretion. Enzymes for digestion of triacylglycerols, phospholipids,
cholesterol esters. Localization of synthesis and activation of these enzymes. The role of
phospholipase A2 and C.
Composition of bile
- Bile salts (cholic acid, chenodeoxycholic acid, deoxycholic acid and lithocholic acid)
- Bile pigments (biliverdin)
- Cholesterol
- Phospholipids
- Water
Role:
- When digesting fats, bile acts as an emulsifier to break the large fat globules into
smaller emulsion droplets. The use of this gives active site for the lipase enzymes (fat
digesting).
- Bile helps fat more soluble by binding fat to its hydrophobic tail and binding water to its
hydrophilic head for transportation of fat tissues.
- Activator of pancreatic lipase
Classification
- Primary bile acids: produced in liver (cholic acid and chenodeoxycholic acid )
+ Synthesized in liver – pass into bile – stored and accumulate in gallbladder.
+ Gallbladder contains 4-5% of bile acids.
+ passed from gallbladder into duodenum for 1st step digestion of lipid
- Secondary bile acids: (deoxycholic acid and lithocholic acid.)
+ Products of transformation of primary bile acid by enzyme of intestinal germs.
Both primary and secondary bile acids are deconjugated in small intestine
Bile is reabsorbed by intestinal mucosa tunica and returned into liver by enterohepatic
circulation in the complex with serum albumin.
Disorder of bile secretion: disorder of secretion bile into enterohepatic circulation.
Bile acids act as main pathway of breaking down and eliminating cholesterol from the
body. The failure in producing or secreting bile acids results in accumulation of abnormal
bile acids and other metabolites, and the excess amount of cholesterol in the body can
damage certain organ system
- Manifestations: malabsorption of fat and fat-soluble vitamin (cannot absorb),
jaundice, abdominal pain (upper right side of abdomen under rib cage), nausea,
vomiting, fatigue, loss of appetite, loss weight
- Diagnosis: blood test, liver function test
- Treatment: addition of missing bile acids
Digestion of TAG, cholesterol esters, phospholipids: Lipids are chemically break down in
interstine. After emulsified by bile in duodenum, fats undergo catalyzed of lipase,
phospholipase, and cholesterol esterase.
TAG:
- Protein colipase is secreted to small intestine, under restrict proteolysis, can activate
active site of lipase. Prolipase (precursor of lipase) is synthesized in pancreas and be
activated by bile acids and protein colipase into lipase
- Activation: Ca2+
- Different kinds of lipase: hormone-sensitive lipase/ TAG lipase, monoacylglycerol lipase
Cholesterol ester:
- Small intestine: enzymes cholesterolesterase catalyzes reaction producing cholesterol
and free fatty acid
- Activation: bisphenol A diglycidyl ether ethanol, phosphatidylcholine methanol,
phosphatidylethanolamine n-butanol, cAMP-dependent protein kinase, type II, sodium
taurocholic acid
Phospholipid:
- In intestine: phospholipase A, A2, C, D catalyze reaction producing lysolecithin and free
fatty acids
+ Phospholipase A2 cut ester bond in 2nd position of glycerol between glycerol and
fatty acid
+ Phospholipase C cut bond between phosphate group and glycerol
- Activation: Ca2+, ATP, Mg2+ (A2)
4. Disorders of digestion and absorption of dietary lipids. Steatorrhea. Vitamin
deficiencies associated with steatorrhea. Features of digestion of lipids in children.
Disorders of lipid digestion and absorption. Steatorrhea
• Absorption:
- The disable of lipid absorption can lead to increase in plasma lipid concentration in
blood, which related to cardiovascular diseases.
• Digestion:
- The body is unable to produce enzyme which necessary to break down lipid
molecules.
- This can lead to accumulation of fat in human body and tissues. Overtime, fats can
damage the brain, peripheral nervous system, liver, spleen and bone marrow
- Eg: Hunter disease (iduronate sulfatase), Hurler disease (alpha L iduronidase), Gaucher
disease (acid Beta glucosidase)
• Malabsorption: disable of unable to absorb nutrients from diet
- Causes: disable of synthesis and secrete required enzyme to break down food, celiac
disease (inflammation in intestine, make it unable to absorb nutrients)
- Symptoms: Brain fog, bloating and nausea after eating, hormonal imbalances, fatigue,
difficulty losing weight or unexplained weight-loss, issues with memory or attention,
-
dry skin and hair, manifestation of absence of fat-soluble vitamin A D E K, steatorrhea
(more than 5% of fat in feces) and streatorrhea
Treatment: addition of required enzymes, treat the inflammation of intestine
• Steatorrhea: presence of excess fat in feces.
- Causes: disease affecting intestine (celiac disease, giardiasis, HIV), deficiency bile
acids, exocrine pancreatic insufficiency (lacks enzyme to digest food)
- Symptoms: bulky, pale, foul-smelling oily stools
- Diagnosis: stool analysis
Prolonged steatorrhea can result in deficiencies of fat-soluble vitamins A, E, D, K, F
Digestion of lipids in children
- Fat synthesis is the most intense in children
- Fat formed from carbohydrates
- In condition lack carbohydrate fat splitting is accompanied by formation of ketone
bodies
5. Resynthesis of lipids in enterocytes. Reactions of resynthesis of triacylglycerols,
cholesterol esters and phospholipids in the intestinal wall. Transport of resynthesized
triacylglycerols in the body.
Resynthesis of triacylglycerols in enterocytes
- Occur after absorption of fatty acids and monoglycerides through villi in small
intestine to enterocytes and converted to triacylglycerols.
- The triglycerides, cholesterol, proteins, phospholipids are packed into chylomicrons
(lipoprotein) by Golgi apparatus.
- The chylomicrons leave the absorptive cell via exocytosis, enter the lymphatic
vessels and transport them to the visceral organs, muscle tissues, adipose tissues.
Resynthesis of TAG, cholesterol esters and phospholipids
TAG
Cholesterol ester:
cholesterol + fatty acyl CoA ---acyl CoA cholesterol acyl transferase--> cholesterol ester +
CoA
Phospholipids
Lysophospholipid + Acyl CoA --lysophosphate acyl CoA acyl transferase--> phospholipid
Transport of resynthesized TAG:
- Blood lipoprotein (chylomicron – contains highest level of TAG)
- Central part is lipid core (TAG, cholesterol esters) and surrounded by single layer
composed of amphipathic lipids (phospholipid, glycolipid, free cholesterol)
- Blood lipoprotein can move within the bloodstream
6. Fatty acid synthesis from glucose: localization and conditions, scheme of acetylCoA
formation from glucose, role of citrate in the transfer of the acetyl group into cytosol,
synthesis of malonyl-CoA, role of biotin. Structure of fatty acid synthase, reactions, the
final product of the synthesis, regulation of the process.
Fatty acid synthesis from glucose:
GLUCOSE -glycolysis-> PYRUVATE -transition-> ACETYL CoA -polymeration -> FATTY ACID
Localization in cytosol and in hyperglycemia condition, insulin stimulation.
Acetyl CoA formed from glucose by glycolysis. Citrate shuttle:
Citrate can cross mitochondria.
Once in cytosol, it is broken down
to reform acetyl CoA and OAA
(catalyzed by citrate lyase). OAA
then transported back to
mitochondria in the form of
pyruvate
Fatty acid synthesis from Acetyl CoA
Biotin acts as cofactor of acetyl CoA carboxylase
which catalyze reaction forming malonyl CoA
from acetyl CoA. This reaction uses 1 ATP
Fatty acid synthase complex: contain acyl carrier
protein (ACP) in the center. ACP contains 4phosphopantothene as prosthetic group.
Besides, it has AT: Acetyl CoA-ACP transacetylase,
MT: malonyl CoA ACP transacetylase, KS: 3
ketoacyl-ACP synthase, KR: 3-ketoacyl ACP
reductase, HD: 3 hydroxyacyl ACP dehydratase,
ER: enoyl-ACP reductase.
Reactions: 4 steps
2 acetyl CoA transformation into malonyl CoA (catalyzed by acetyl CoA carboxylase,
required biotin as cofactor) and then four steps of FAS to lengthen the chain by 2 carbons,
and another malonyl group is linked to the chain. After 7 cycles of condensation and
reduction produce 16-carbon saturated palmitoyl group.
- Step 1 (condensation): activated acetyl and malonyl groups condensate with
formation of acetoacetyl-ACP (acyl carrier protein)
- Step 2 (reduction of carbonyl group): acetoacetyl-ACP ---reduction---> 3hydroxybutyryl-ACP
- Step 3 (dehydration): dehydration of 3-hydroxybutyryl-ACP to yield double bond in
product trans-enoyl-ACP
- Step 4 (reduction of double bond): saturate double bond of trans-enoyl-ACP --enoylACP reductase--> butyryl-ACP
- 1 pass through fatty acid synthase complex forms 4-carbon saturated fatty acyl-ACP.
Butyryl group is transferred from phosphopantothene-SH group to Cys-SH- group of 3
ketoacyl-ACP synthase.
- Another malonyl group is linked to the chain for repeated reactions on FAS
- 7 cycles of condensation and reduction produce 16-carbon saturated palmitoyl groupACP.
- Hydrolysis release free palmitate from ACP
Regulation:
- Insulin activates transformation of citrate to acetyl CoA
- Citrate activates transformation of CoA to Malonyl CoA, while glucagon, epinephrine
inhibits this reaction
- Palmitoyl CoA inhibits the reaction forming it from malonyl CoA
7. Synthesis of glycerol 3-phosphate from glucose. Localization and biologic role of the
process. Synthesis of phosphatidic acid from glycerol 3-phosphate and fatty acids:
localization in the cell, the sources of glycerol-3-phosphate, fatty acids and energy, the
sequence of reactions, interconnection with carbohydrate metabolism, metabolic
pathways for phosphatidic acid.
Synthesis of glycerol 3 phosphate: in cytoplasm of cell to produce metabolite of
lipogenesis, gluconeogenesis, and glycolysis
Synthesis of phosphatidic acid from glycerol 3P and fatty acid: 2 pathways:
- Phosphatidic acid is formed by addition of 2 fatty acyl CoA to glycerol 3P and release
CoA-SH
- Phosphorylation of diacylglycerol by diacylglycerol-kinase (using 1 ATP)
Interconnection with carbohydrate metabolism:
DHAP: Glycolysis, gluconeogenesis intermediate
Glycerol 3P: gluconeogenesis intermediate
Phosphatidic metabolic pathway:
8. Reactions of synthesis of triacylglycerols (lipogenesis). The fatty acid composition of
triacylglycerols. Interconnection with carbohydrate metabolism. Synthesis of
triacylglycerol in adipose tissue and in liver.
TAG is synthesized from fatty acyl CoA and glycerol 3P
- In main tissue, glycerol 3P is formed from glycolytic
intermediate DHAP under catalyzed of G3P DH
- In the liver, glycerol –glycerol kinase--> G3P
- Fatty acid ---acyl CoA synthase ---> Fatty acyl CoA
- Acylation of glycerol 3 phosphate by fatty acid CoA yielding phosphatidic acid under
catalyzed of acyl CoA transferase
- Phosphatidic acid is hydrolyzed by phosphatase to yield DAG
- Acylation of DAG by fatty acid CoA yielding TAG under catalyzed of diacylglycerol CoA
transferase
Fatty acids component of TAG
- Saturated: Stearic acid and palmitic acid
- Unsaturated: α-linolenic acid (ALA), linoleic acid (LA), arachidonic acid and gamma
linoleic acid
Interaction between lipid and carbohydrate metabolism
- Pyruvate end product of glycolysis is oxidative decarboxylated to acetyl CoA
- Acetyl CoA is the link between both pathways
- Glucose, glycerol and fatty acids are degraded into acetyl CoA
- Glyceraldehyde 3 phosphate is an intermediate for glycolysis and source of glycerol
synthesis
- Biosynthesis of fatty acids, ketone bodies and cholesterol all use acetyl CoA
- Free excess glucose is using as source for lipogenesis
Synthesis of TAG in liver and adipose tissue:
- In both liver and adipose tissue, glucose --> DHAP ---> glycerol 3 phosphate
- In only liver, glycerol kinase convert free glycerol to glycerol phosphate
9. Lipolysis: localization and conditions, a sequence of reactions and enzymes, the final
products, hormonal regulation, transport of produced fatty acids and their using,
utilization of glycerol. The energy effect of glycerol oxidation.
Lipolysis: occurs in adipose tissue, under influence of glucagon/epinephrine/
norepinephrine when blood glucose is low
Reactions and enzyme
- Hydrolysis of lipids forming free fatty acids and glycerol by adipose cell lipase
- Glycerol is phosphorylated to glycerol 3P and then DHAP which in turn isomerized to
glyceraldehyde 3 phosphate (glycolysis intermediate of glycolysis, gluconeogenesis)
- Fatty acids bind to albumin and transported to tissue for energy production.
Hormonal regulation
- (+) hypoglycemia: glucagon, epinephrine, norepinephrine
- (-) hyperglycemia: insulin
Glycerol oxidation: forming intermediate of glycolysis and gluconeogenesis to regulate
blood glucose level
10. Reactions of fatty acid oxidation to carbon dioxide and water: the role of carnitine,
localization and conditions, reactions of β-oxidation, role of vitamins and coenzymes,
the final products, connection with the TCA cycle and respiratory chain, the energy yield
of the process, the calculation of the energy effect of ß-oxidation of palmitic acid.
Fatty acid oxidation: is 4-step pathway to produce acetyl CoA essential for citric acid
cycle and oxidative phosphorylation to produce ATP, release CO2 and H2O
- In cytoplasm, fatty acids are acylated by acyl CoA synthetase forming fatty acid CoA
Fatty acid + CoA-SH + ATP --acyl CoA synthetase--> Fatty acid S-CoA + AMP + H4P2O7
- Carnitine shuttle transfers fatty acid CoA from cytosol to mitochondria matrix
+ CAT1 (carnitine acyl transferase- located in outer surface of mitochondria) adds
carnitine to fatty acid CoA and releases CoA-SH, forming fatty acid carnitine which can
get to mitochondria matrix
+ CAT2 (located in inner surface of mitochondria) transfer carnitine back to cytosol and
adds CoA-SH to fatty acid, forming fatty acid CoA for beta oxidation
- Beta oxidation of fatty acid CoA
Products: from Palmitoyl CoA (16C) form:
- 8 acetyl CoA - 7 FADH2 - 7 NADH + 7 H+
All products enter citric acid cycle and oxidative
phosphorylation
After citric acid cycle: 1 acetyl CoA -> 1 ATP, 3 NADH,
1 FADH2
After oxidative phosphorylation: 1 NADH -> 3 ATP; 1
FADH2 -> 2 ATP
1 acetyl CoA --> 12 ATP
Total forming ATP: 8x12 + 7x2 + 7x3 = 131 ATP
ATP utilize: 1 ATP
TOTAL PRODUCT: 130 ATP
Vitamin and coenzyme:
FAD (coenzyme for Acyl CoA DH) and NAD+ (b-hydroxyacyl CoA DH)
11. Triacylglycerol metabolism in different physiologic states (food intake, fasting,
muscle activity).
- During food intake blood glucose level is high (hyperglycemia). The excess amount of
glucose can we use for lipogenesis. This process is influenced by insulin
- During fasting or muscle activity, blood glucose level decreases (hypoglycemia).
Glycerol, fatty acids from lipolysis are taken for glucogenesis or glycolysis. Glucagon or
epinephrine stimulate these processes
12. Reactions of the synthesis of ketone bodies. Conditions, localization, biologic role.
Reactions of ketone bodies utilization in tissues.
Ketogenesis: in hypoglycemia and glucose is not available, body uses fatty acids as
source fuel for synthesize acetyl CoA through forming ketone bodies. This process takes
place in the liver and ketone bodies can move to muscle through bloodstream.
Role: when the glucose in human body is not available for body function, fatty acids are
used to form acetyl CoA which is entrance substrate of citric acid cycle to generate ATP
for brain and muscle activities.
Ketone utilization:
Acetyl CoA then goes to citric acid cycle and produce
energy
13. Ketoacidosis in starvation and in diabetes mellitus. Role of oxaloacetate deficit in
activation of ketogenesis.
Ketoacidosis in starvation and in diabetes mellitus.
- In untreated diabetes mellitus (type I) (wo insulin, body use fat as fuel ) and prolonged
starvation, glucose blood level is low (hypoglycemia). Fatty acid is used as fuel for brain
and muscle function by forming ketone bodies when glucose is not available
- Ketone bodies (acetoacetate, b-hydroxyl butyrate) which are synthesized in the liver
get to muscle, brain through bloodstream are acidic. They decrease blood pH -->
ketoacidosis
Role of OAA deficit
- Deficit of OAA (caused by decrease of glucose -> pyruvate -> acetyl CoA) decreases rate
of citric acid cycle which then decrease energy synthesis. It stimulates the body to use
secondary fuel which is fatty acids --> KETOGENESIS
14. The fatty acid composition of phospholipids. Reactions of phospholipids biosynthesis
in tissues. Two pathways of phospholipid biosynthesis. The role of vitamins B6, B9 and
B12, methionine and serine. Disorders of phospholipid synthesis. Hepatic steatosis (fatty
liver).
Fatty acid composition of phospholipids: saturated and unsaturated fatty acid.
Phospholipid biosynthesis: 2 pathways
- Synthesized from phosphatidic acid (produced from glycerol 3 P) and diacylglycerol
(intermediate of TAG synthesis)
- Synthesis from ethanolamine
Ethanolamine + ATP --ethanolamine kinase--> phospho-ethanolamine + ADP
Phospho-ethanolamine + CTP (cytidine triphosphate) --> CDP ethanolamine + H4P2O7
DAG + CDP ethanolamine --> phosphatidyl ethanolamine + CMP
Phosphatidyl ethanolamine + 3 SAM (S-adenyl methionine) –methyltransferase-->
phosphatidyl choline
There is interconnection pathway between phosphatidyl choline, phosphatidyl serine
and phosphatidyl ethanolamine
Role of vitamin B6, B9, B12, methionine and serine
- B6: cofactor in enzyme involved in transamination decarboxylation sphingolipid
synthesis
- B9: carry and transfer one carbon unit during biosynthesis reactions
- B12: cofactor in enzymes of conversion malonyl CoA to succinyl CoA and conversion of
homocysteine to methionine
- Methionine: under transferation forming SAM which is essential for phosphatidyl
choline biosynthesis
- Serine: required for convert of phosphatidyl ethanolamine to phosphatidylserine
Disorder of phospholipids synthesis: deficiency in factors of phospholipid biosynthesis
- B12 deficiency -> megaloblastic anemia
- B9 deficiency -> disable of mitosis in DNA synthesis
- B6 deficiency -> damage to nervous system by disorder of sphingomyelin synthesis
- SAM -> disable converting phosphatidyl ethanolamine to phosphatidyl choline for
membrane component
- Serin -> disable forming phosphatidyl serine for cell membrane component
Hepatic steatosis (fatty liver): excess fat builds up in the liver
- Causes: diabetes mellitus, alcohol, obesity, drugs, celiac disease, glycogen storage
diseases,
- Manifestation: no or few symptoms, maybe tiredness or pain in abdomen
- Complications may include cirrhosis, liver cancer, and esophageal varices.
- Diagnosis: liver function test
15. Lipidosis: Tay-Sachs disease, Gauchers disease, Niemann-Pick diseases.
Tay Sachs disease
Causes: disorder of deficiency of hexosaminidase A which is involved in the breakdown
of gangliosides (glycolipid contains neuraminic acid)
Manifestation: loss of motor skill, cherry red-spot on retinal examination, increased
startle reaction to noise
Biochemical aspect: deficiency of enzyme leads to accumulation of GM2 gangliosides in
ganglion cells of CNS which are toxic to neuronal cells and lead to neurological damage
Treatment: symptom supportive treatment
Gauchers disease
Causes: disorder deficiency of acid beta glucosidase
Manifestation:
- Type I (in adult): seizure, aseptic necrosis
- Type II (in infant): mental retarded
- Type III (juvenile): dementia
Biochemical aspect: accumulation of glycosphingolipid in brain, liver, spleen and bone
marrow cause destruction to organ
Diagnosis: bone marrow biopsy
Treatment: cerezyme which is synthetic form of deficiency enzyme
Niemann Pick disease
Causes: disorder resulted deficiency of sphingomyelinase
Manifestation: seizures, atazia, dystonia, mental retardation, intertitial pulmonary
disease, osteoporosis, pulmonary hypertension, abdominal, spleen or liver enlargement
Biochemical aspect: sphingomyelin accumulate in lysosomes of brain, liver, spleen,
bone marrow and lungs lead to dysfunction of these organs
Diagnosis: family history, genetic testing
Treatment: bone marrow transplantation, enzyme therapy
16. The chemical structure and biological role of cholesterol. Dietary sources of
cholesterol. Pathways of cholesterol metabolism. Removing cholesterol from the body.
Cholesterol
Structure: consists of four linked hydrocarbon rings forming
the bulky steroid structure and hydrocarbon tail linked to one
end of the steroid and a hydroxyl group linked to the other
end
Biological role: precursor of bile acids, steroid hormone and
vitamin D; formation of myelin sheath; stabilize and provide
less fluidity and prevention of phase transition of cell
membrane.
Dietary sources: egg yolk, shrimp, beef, cheese, butter
Cholesterol metabolism:
- Cholesterol synthesis - Bile acid synthesis - Steroid hormone synthesis - Vitamin D
synthesis - Cholesterol ester synthesis
Cholesterol is removed from the body: under bile salt form, through enterohepatic
circulation
17. Cholesterol biosynthesis. Localization, substrates, stages. Reactions for mevalonate
synthesis. Scheme of the next stages. Interaction with carbohydrate metabolism.
Regulation of cholesterol synthesis. Hormonal and allosteric regulation mechanisms.
Drug regulation of cholesterol synthesis.
Cholesterol biosynthesis: using Acetyl CoA in the liver to produce cholesterol
Stages: 4 stages
1. Synthesis of mevalonate
2. Conversion of mevalonate to active isoprene unit
Phosphorylation of mevalonate
forming isopento diphosphate
accompanied with using ATP
Isopentyl diphosphate and
dimethylallyl diphosphate
isomerization forming active
isoprene unit
3. Polymerization of 6 isoprene units form squalene (30 carbons)
4. Cyclization of squalene yield 4 rings of steroid nucleus of cholesterol.
Interaction with carbohydrate metabolism:
- Pyruvate end product of glycolysis is oxidative decarboxylated to acetyl CoA
- Acetyl CoA is the link between both pathways
Regulation of cholesterol synthesis:
- Hormonal: HMG CoA reductase is stimulated by insulin and inhibited by glycagon
- Allosteric: high level of cholesterol inhibits HMG CoA
Drug: HMG CoA is regulated by lovastatin to reduce cholesterol synthesis
18. Bile acids, classification, structure and physiological role. Stages of the synthesis of
bile acids, Role of vitamins. Primary and secondary bile acids, conjugated bile acids.
Enterohepatic circulation of bile acids.
Bile acids: produced from cholesterol in the liver and it emulsifies fats (make it soluble)
and activates lipase for digestion
Classification
- Primary bile acids: produced in liver (cholic acid and chenodeoxycholic acid )
+ Synthesized in liver – pass into bile – stored and accumulate in gallbladder.
+ Gallbladder contains 4-5% of bile acids.
+ Passed from gallbladder into duodenum for 1st step digestion of lipid
-Secondary bile acids: (deoxycholic acid and lithocholic acid.)
+ Products of transformation of primary bile acid by enzyme of intestinal germs.
Synthesis of bile:
- Primary bile acids are synthesized in liver by transformation of cholesterol. They pass
into the bile and then stored and accumulated in gallbladder.
- Bile passes from gallbladder into duodenum. Secondary bile acids are products of
transformation of primary bile acids by enzyme of intestinal germs.
Vitamin
- Fat soluble vitamin (A and D) inhibits bile acid synthesis by repress rate-limiting
enzyme CYP7A1 which catalyzes cholesterol transformation.
Conjugated bile acids: conjugation between glycine or taurine wit bile acids. (8 forms:
primary bile acids)
- Glycocholic acid is a major conjugated bile acid in the bile
- Conjugated bile acids are present in bile as sodium salts
Enterohepatic circulation: the process which these 4 bile acids transform to each other
and recycled. This process is important in toxicology and lipophilic xenobiotics.
-
Synthesis of bile salts and conjugation
-
Conjugated bile salts go from liver to small intestine
-
Expenditure of bile salt in small intestine and excretion of bile acids in feces (5%)
-
Deconjugation of bile salts through portal vein (95%)
19. Disorder of cholesterol metabolism and gallstone disease. Role of phospholipids.
Lipotropic factors.
Disorder of cholesterol metabolism: defect in enzyme necessary for cholesterol
metabolism
- Happle syndrome: high cholesterol level leads to caratact, skeletal dysplasia
- Child syndrome: mutation in cholesterol biosynthesis enzyme
- Ichthyosis:
Gallstone disease: excess cholesterol can form crystal-like stone in gallbladder called
gallstone.
Role of phospholipids:
- Component of cell membrane
- Form the structure of fat and transported it throughout the bloodstream
- Storage energy
Lipotropic factors: are produced naturally in the body. They are substances that can
remove and prevent fatty deposits, such as Homocysteine, in the body.
20. Lipoproteins of blood: classification, structure, stages of formation. Apoprotein:
classification, function.
Lipoproteins are globular particles which consist of lipid core (TAG, cholesterol ester)
and surrounded single layer sheet (phospholipid, glycolipids, free cholesterol) binding
proteins by noncovalent bonds.
Classification:
According to their density: chylomicron, VLDL, LDL, IDL, HDL
According to their electrophoretic mobility depends on content of apoprotein in its
structure
Stages of formation:
- Lipids after absorption is resynthesized.
- Nascent lipoprotein is released to the blood and accepts apoprotein to become mature
- Mature lipoprotein reaches target tissue (muscle, adipose tissues) and be hydrolyzed
by lipase
Apoprotein: protein part of lipoprotein
- Function: recognition molecules for membrane receptors; enzymes that are involved in
lipid metabolism; coenzyme of lipid metabolism enzymes.
- Classification: A1, A2, A4, B48, B100, C1, C2, C3, D and E
21. Transport of dietary triacylglycerol in organism. Chylomicrons: lipid composition,
structure, functions, apoproteins. Localization of chylomicron formation. Role of
lipoprotein lipase in chylomicron metabolism.
Transport of dietary TAG in organism mainly by chylomicrons
Chylomicrons
- Lipid composition: free cholesterol (1-3); cholesterol ester (3-5); phospholipid (7-9);
TAG (84-89)
- Structure: 100-1000nm in diameter
- Function: transfer absorbed TAG in intestine to other tissues
- Apoprotein: apoprotein A, B48, B100, C, E
- Localization of formation: tunica mucosa of intestine
- Role of lipoprotein lipase: forming free fatty acids, glycerol for cell utilization
22. Sources of TAG in the liver. Very low density lipoproteins: lipid composition,
structure, functions, apoproteins. Role of lipoprotein lipase in VLDL metabolism.
Sources of TAG in the liver: VLDL
Very low density lipoprotein
- Lipid composition: free cholesterol (5-10); cholesterol ester (10-12); phospholipid (1520); TAG (50-65)
- Structure: 25-75nm in diameter
- Function: transfer TAG from hepatocyte released to the blood and be uptake by the
liver
- Apoprotein: apoprotein C, E
- Localization of formation: hepatocyte
- Role of lipoprotein lipase: forming free fatty acids, glycerol for hepatocyte and free
circulate in the blood for tissue absorbtion
23. Localization and role of apo B100 receptor. Role of receptor-mediated endocytosis
of LDL, pathway of components after endocytosis. Role acyl SCoA: cholesterol
acyltransferase (ACAT).
Apo B100: located in extrahepatic tissues, acts as receptor of LDL to the liver
Role of receptor mediated endocytosis of LDL (apo B100): allows the invagination of LDL
and forms endocytic vesicle. This vesicle later fuse with lysosome in the liver
Pathway of components after endocytosis:
- All components are released to liver cell
- Cholesterol and fatty acids are resynthesized for membrane synthesis, steroid
hormone synthesis, vitamin D synthesis and bile acid synthesis
Cholesterol acyltransferase (ACAT) catalyzes esterification of cholesterol and free fatty
acid in tissue for synthesis of steroid hormones, vitamin D, bile acids and membrane.
24. Transport of cholesterol and cholesterol esters in blood. Role of high density
lipoproteins and low density lipoproteins. Lecithin-cholesterol acyltransferase reaction.
Transport of cholesterol and cholesterol esters in the blood by HDL and LDL
Role of HDL and LDL
- HDL: remove cholesterol from cholesterol-laden cells and return it to the liver
- LDL: transfer free cholesterol and cholesterol ester to liver – main site of cholesterol
utilization
Lecithin-cholesterol acyltransferase reaction (activator: apoAI) converts free
cholesterol to cholesterol esters and lysolecthin. Cholesterol ester move into
hydrophobic part of bilayer and lysolecthin is combined to plasma albumin
25. Role of essential polyunsaturated fatty acids in cholesterol metabolism. Vitamin F:
dietary sources, daily requirements, biochemical functions, signs of deficiency.
Polyunsaturated fatty acids ω3: examples, structure, properties, biologic role.
Eicosanoids: prostaglandins, thromboxanes, leucotriens. Biologic role. Scheme of
synthesis. Role of phospholipase A2, cyclooxygenase, lipooxygenase. Factors affecting
the synthesis of eicosanoids.
Role of essential polyunsaturated fatty acids in cholesterol metabolism:
- Polyunsaturated fatty acids may lower serum very low-density and low-density
lipoprotein concentrations because the liver preferentially converts polyunsaturated
fatty acids into ketone bodies
Vitamin F:
- Complex of polygenic fatty acids (polyunsaturated fatty acids): alpha linolenic acid
(ALA) and linoleic acid (LA
- Sources: plant oil, fish, milk, dairy product
- Significance: membrane phospholipid (cell membrane component); increase
polyunsaturated fatty acid -> increase properties of cell membrane; precursor of some
active substances in human body (bile)
- Hypoviaminosis: disorder genetic factor, reproduction factor, atherosclerosis
Polyunsaturated fatty acids omega 3
Eg: α-linolenic acid (ALA), found in plant oils, and eicosapentaenoic acid (EPA),
docosahexaenoic acid (DHA) in marine oils and docosapentaenoic acid (DPA)
Structure: contains double bond, three atoms away from the terminal methyl group in
their chemical structure
Properties:
- ALA is converted to EPA or DPA by chain elongation and desaturation
- Forming esters with cholesterol to decrease LDL and VLDL
Role:
- Required for normal growth especially in children
- Main component of brain grey matter, retina, testis, sperm
- Reduce cardiovascular disease
Eicosanoids: are signaling molecules made by the enzymatic or non-enzymatic oxidation
of arachidonic acid or other polyunsaturated fatty acids (PUFAs)
- Prostaglandins: vasodilator, autocrine or paracrine factors
- Thromboxanes: vasoconstrictor, facilitate platelet aggregation
- Leucotriens: autocrine or paracrine factors
Biological role
- Act as hormone to regulate cell activities,
- Involved platelet aggregation
- Constriction, dilation for blood vessel and bronchi
Synthesis
Phospholipase a2: catalyze hydrolysis of phospholipid/DAG to release arachidonic acid
Cyclooxygenase: Conversion of arachidonic acid to prostaglandin, thromboxane
Lipoxygenase: catalyze conversion of arachidonic acid to leukotriene synthesis
intermediate
26. Hyperlipoproteinemia type IIA (familial hypercholesterolemia): mechanisms and
clinical simptoms.
Mechanism: defect of 4 classes of LDL leads to reduce LDL clearance ->
hypercholesterolemia
Clinical symptoms: coronary artery disease, atherosclerosis -> pain, fatigue, muscle
weakness
27. Atherosclerosis: stages. The role of modified LDL in the initiation of atherosclerosis.
Neutrophils and monocytes in pathogenesis of atherosclerosis.
Atherosclerosis:
Stages
1. Endothelial dysfunction
2. Formation of lipid layer or fatty streak within the intima
3. Migration of leukocytes and smooth muscle cells into the vessel walls
4. Foam cell formation
5. Degradation of extracellular matrix
Role of modified LDL in the initiation of atherosclerosis
- When the epithelial cells are damage, it starts secreting adhesive molecules to bind
circulating monocytes. Accumulation of monocyte are transformed into macrophages
which can express as receptor for oxidatively modified LDL -> forming foam cells
- Foam cells accumulate underlying extracellular matrix of the epithelial cells. These
exposed areas serve as site for platelet adhesion and aggregation.
- Platelets secrete cytokines which promote formation of thrombus -> atherosclerosis.
28. Characteristics of disorders of triacylglycerols transport - dyslipoproteinemia types I
and V. mechanisms and clinical symptoms.
Dyslipoproteinemia: defect in lipoprotein metabolism leads to hypo or
hyperlipoproteinemia
Dyslipoproteinemia type I: HYPERlipoproteinemia
- Mechanism: deficiency of LPL or apoC lead to slow clearance of chylomicron and
increase TAG level in the blood.
- Symptoms: abdominal pain, diabetes
Dyslipoproteinemia type V: HYPERlipoproteinemia
- Mechanism: elevated level of chylomicrons and VLDL leads to increasing TAG and
cholesterol with decrease of HDL
- Symptoms: fatigue, abdominal pain, pancreatitis, enlarge liver or spleen, xanthomas
MODULE V. NITROGEN METABOLISM
1. Nitrogen balance. Negative, positive nitrogen balance. Equilibrium. Physiologic and
pathologic situations with different nitrogen balance. Nitrogen balance in children. Food
sources of proteins. Daily requirement for proteins in adults and in children. The
biological value of proteins. Manifestations of protein deficiency. Kwashiorkor.
Nitrogen balance = nitrogen intake – nitrogen output
(Nitrogen containing substances: aa, nucleic acid))
- Negative nitrogen balance: intake is less than output. Causing chronic/acute illness,
protein deficiency, starvation, stress, senescence, sepsis, fever, kwashiorkor.
Hormones lead to NNB: corticosteroids, T4
- Positive nitrogen balance: intake is more than output. Bringing back growth,
pregnancy, recovery from illness. Hormones lead to PNB: growth hormone, insulin,
androgens
- Nitrogen equilibrium: intake = output: normal condition
In children, nitrogen balance maintaining in positive balance (1.7g protein/kg/day) is
necessary for their growth.
Dietary proteins: from meat, dairy products, nuts, certain grains and beans
Daily requirement: In child, intake protein requirement is 1.7g protein/kg/day to maintain
positive nitrogen balance and in adult is 0.8g protein/kg/day. In Russia, sum requirement
of protein of an adult is 80-115g.
Nutritional value: 1 gram provides 17kJ (4kcal) (high value in animal protein and low value
in plant protein)
Manifestation of protein deficiency:
- Skin, hair and nail problems
- Loss of muscle mass
- Brain, kidney injury
- Increasing risk of bone fractures
- Risk of infections
- Fatty liver
- Bigger appetite and increased calories intake
By the age of 7, if protein deficiency is not corrected by suitable diet in time can lead
to dead
Kwashiorkor: Kwashiorkor, also known as “edematous malnutrition” because of its
association with edema (fluid retention), is a nutritional disorder with liver enlargement
- Cause: lack of protein in the diet.
-
-
Manifestations: People who have kwashiorkor typically have an extremely emaciated
appearance in all body parts except their ankles, feet, and belly, which swell with fluid.
Symptoms: change in skin and hair color (rust color), fatigue, diarrhea, loss of muscle
mass, failure to grow or gain weight, edema of ankles, feet and belly, damage immune
system, …
Treatment: increase protein intake
2. Digesting of proteins in the stomach and in intestine. The mechanism of the synthesis
and the biological role of gastric acid. Disorders of gastric acid production. Enzymes of
gastric juice, pancreatic juice and intestinal juice involved in the digestion of proteins.
Digestion of proteins in stomach:
- Begins in stomach with secretion of gastric juice which contains hydrochloric acid and
pepsin by cells of stomach mucosa tunica.
- Hydrochloric acid actives pepsinogen into pepsin.
- Pepsin (endopeptidase) hydrolyzes only peptide bond formed by aromatic amino acids
(tyrosine, phenylalanine) or aspartate and glutamate, forming shorter peptides
compare to initial peptides and only some free amino acids.
- Besides, gastrixin are synthesized in chief cells of stomach mucosa tunica. In stomach
of newborn, chymosin is synthesized (proenzyme: prochymosin) to hydrolysis milk
protein which contains casein majorly.
Digestion of proteins in intestine
- Peptide together with hydrochloride acid move to duodenum. Peptides stimulate the
secretion of cholecystokinin which is a hormone peptide stimulating secretion of
pancreatic proteases. HCl stimulates secretion of pancreatic juice to neutralize pH by
carbonic acid to create optimal pH for pancreatic proteases
- All pancreatic pro-enzyme are secreted by exocrine part of pancreas to lumen of
duodenum and transformed into active enzymes. (PP: trypsin, chymotrypsin,
carboxypeptidase A. collagenase, elastase)
Parietal digestion: (terminal intestine) hydrolyzation of short peptide into free amino
acids by aminopeptidase. Aminopeptidase is secreted by microvilli of small intestine and
it is zinc dependent enzyme
Gastric acid:
Role:
- Process of protein denaturation in stomach
- Activation of pepsin accompanied by transformation of inactive pepsinogen to pepsin
- Formation of pH of gastric juice equalizing pH optimum of pepsin
-
Disinfection of stomach content
Synthesis:
- Carbon dioxide released from citric cycle in mitochondria goes to cytosol. Carbon
dioxide reacts with water forming carbonic acid.
CO2 + H2O ↔ H2CO3
Carbonic anhydrase
H2CO3 ↔ HCO3- + H+
- Carbonic acid undergoes dissociation to yield H+ proton.
- H+ is transferred by membrane enzyme H+ K+ - ATPase (H+ out, K+ in)
- Cl- passes from cell into gastric juice (by Cl- channel), together with H+ forming HCl in
gastric juice
Regulation:
- (+) ACh (vagus nerve), Histamine (mast cells), gastrin (G cells)
- (-) Acid, somatostatin
Disorder of HCl production:
* hypochlorhydria: unable to produce enough HCl in stomach
- Causes: aging, stress, medications (long term use of antacids), bacterial infection, zinc
deficiency, stomach surgery.
- Symptoms: bloating, burping, diarrhea, hair loss, heartburn, intestinal infections,
nausea while taking supplements, undigested food in stool, upset stomach.
- Diagnosis: acidity of stomach test
- Treatment: antibiotics for H.pylori (for intestinal infection), get rid of antacids
medication, digestive enzymes and HCl supplements, adding nutrient deficiencies
(zinc)
* Hyperchlorhydria: over production of HCl in stomach
* Achylia: absence of both HCl and pepsin in gastric juice (total achylia leads to lack of
intrinsic factors which is used to absorb vitamin B12)
Gastric juice: pH: 0.7-4
- Chemical composition: hydrochloric acid (produced by parietal cells), pepsin
(produced by chief cells) and mucin (special glycoprotein forming the tunica - Pepsin:
- Highest catalytic effect at pH 1.0-1.5 (produced by HCl)
- Optimum temperature: 37-42C
- Only hydrolyzes peptide bond formed by aromatic amino acids (tyrosine,
phenylalanine) or asparate and glutamate.
- Forming shorter polypeptides.
Pancreatic juice:
- Contain proenzyme of all pancreatic protases. It is stimulated to secreted by appear of
peptides and HCl in duodenum (by action of CCK).
- Components: chymotrypsinogen, procarboxypeptidase, proelastase, procollagenase
- Pancreatic juice helps neutralize the pH of HCl to provide optimal environment for
pancreatic enzymes (by using carbonic anhydrase, forming HCO3-)
Intestinal juice: contains activated pancreatic protases. Those enzymes are activated by
trypsin. And trypsin is activated by HCl.
- Trypsin: Trypsin has pivotal role in activation of pancreas proenzymes. Trypsinogen
(249 aa) which consists of one polypeptide chain undergoes proteolytic by
enterokinase (which is secreted by tunica mucosa of duodenum) forming active
trypsin (243aa) and inactive hexapeptide (6-7 aa).
- Trypsin hydrolyzes peptide bonds between Arg, Lys, His. Trypsin optimum pH is 7.5-8.5
- Trypsin then activates other pancreatic protases by proteolysis
-
Chymotrypsinogen
→
Chymotrypsin
(hydrolyzes peptide bonds between Leu, Trp, Tyr, Phe)
Procarboxypeptidase A
→
Carboxypeptidase A
(hydrolyses C-terminal of peptide chain)
Proelastase
→
Elastase
(hydrolyzes peptide bonds between Ser, Gly,Ala)
Procollagenase
→
Collagenase
(hydrolyzes peptide bonds in collagen)
Polypeptides are cut into short peptide chain
Parietal digestion: hydrolyzation of short peptide into free amino acids by
aminopeptidase. Aminopeptidase is secreted by microvilli of small intestine and it is zinc
dependent enzyme
3. Disorders of protein digestion and absorption, connection of these disorders with the
development of allergic reactions. Features of protein digestion and absorption of amino
acids in children of different ages. Celiac disease.
Disorders of protein digestion and absorption:
- Disorder HCl production
- Disorder in enzyme secretion --> self-digestion: pancreatitis
Connection with development of allergic reactions
- Food protein which cannot be break down circulate in the blood and recognized as
foreign particles. The response of immune system to it cause allergy
Protein digestion and absorption in children require different enzymes due to their food
and HCl amount:
CHYMOSINE:
- Proteolytic enzyme synthesized in newborn stomach. Its secretion is maximal during
first few days after birth and declines after, replaced by pepsin
- Proenzyme: prochymosine, activated by Ca2+ in casein
- Optimum pH: 3.0-4.0
- This enzyme participates in coagulation of milk and hydrolysis of milk proteins (casein).
Casein undergoes of chymosine attack in presence of Ca2+ and absence of HCl in gastric
juice
GASTRIXIN:
- Synthesized as proenzyme in chief cells in stomach mucosa tunica
- Optimal pH: 3.0-3.5
- Amount of gastrixin is many times less than pepsin.
Celiac disease: Pancreatitis: self-digestion of pancreas. It happens when trypsin is
activated not in duodenum lumen. Trypsinogen is not secreted by exocrine part of
pancreas and forming active trypsin. Trypsin actives zymogen and turn pancreatic
proteases on and leads to pancreas self-digestion.
- Manifestations: inflammation of pancreas, focal necrosis
- Treatments: conservative treatment, surgery treatment, medicine (contrical, calcilol)
4. Decomposition of proteins in intestine under influence of microbial enzymes.
Substances formed by the decay of proteins. The processes of detoxification in the liver:
microsomal oxidation and conjugation system. Reactions of indican formation.
Decomposition of proteins in intestine under influence of microbial enzymes:
Microbial flora in large intestine produces enzymes to ferment free amino acids.
Fermented amino acids are food for microorganism to grow and its grown provide
amino acids for human body
Enzymes of microorganisms (in colon or dead organisms) decarboxylate:
- amino acids to form diamine (cadaverine, putrescine)
- from pyruvic acid to form indole (skatole, indole)
- sodium salt of salicyclic acid to form phenol (phenol, cresol)
- amino acid containing sulfur to form hydrogen sulfide or methyl mercaptan.
Detoxification: 2 phases
Microsomal oxidation:
- In the liver, NADPH are taken to the electron transport chain (flavoprotein 1) and then
it will activate the 2nd electron transport carrier cytochrome P450
- cytochrome P450 catalyses the hydroxylation (turn RH into ROH) and make toxic
substance hydrolyse to increase its solubility
- Toxic can cross the membrane, into the blood and eliminated by kidney filtration
Conjugation system: makes the substrate more water soluble and capable of being
excreted by liver or biliary rout or renal route or blood and kidney filtration.
- Glutathione, sulfate, and glucuronic acid serve for conjugation of xenobiotics.
Indican formation
- Following absorption from the gut, indole together with sulfuric acid and potassium
on the decomposition of tryptophan is converted to 3-hydroxy indole (indoxyl or
indican).
- In the liver, where it is again then conjugated with sulfuric acid or glucuronic acid
through normal xenobiotic metabolism pathways. It is then transported to the kidneys
for excretion
5. The transport of amino acids through cell membranes. Sources and pathways of
amino acids in tissues. Glucogenic and ketogenic amino acids.The fate of α-keto acids
formed in the process of deamination (for pyruvate, oxaloacetate, alpha-ketoglutarate).
The calculation of the energy effect of oxidation of the amino acids.
Transport of amino acids through cell membranes: 2 ways
-
Active transport associated with using energy of transmembrane sodium gradient
(catalyzed by Na+ K+ ATPase)
Transport which is provided by glutathione system
Sources and pathways of amino acids in tissues:
Sources: from daily intake food
Fate of free amino acids in small intestine
- 50%: absorbed in small intestine and go to visceral organs by blood stream. This is
exogenous proteins
- 10%: undergoes chemical transduction by bacteria in large intestine
- 40%: pass through large intestine as feces
Glucogenic amino acids are aa that can be converted into glucose through glucogenesis.
It includes most standard amino acids: alanine, aspartate, arginine, cysteine, glutamate,
glutamine, proline, serine, asparagine, histidine, methionine, valine
Ketogenic amino acids are aa that can be converted into acetyl CoA or acetoacetetyl CoA
which are precursor of ketone bodies. It includes lysine and leucine only.
Aa are both glucogenic and ketogenic: tyrosine, isoleucine, phenylalanine, tryptophan,
threonine
Fate of alpha ketoacids formed in the process of deamination
Pyruvate: Pyruvate + CO2 –pyruvate carboxylase→ oxaloactetate
Produce metabolite for citric cycle and gluconeogenesis
- Pyruvate + NAD+ –pyruvate dehydrogenase→ acetyl CoA + NADH + H+ + CO2
Entry citric cycle to produce energy, fatty acid synthesis
• Oxaloacetate: can be used in citric acid cycle and gluconeogenesis
• A-ketoglutarate: involved in transamination in synthesis and catabolism of amino
acids. Used for synthesis of glucose, formation of fat/ketone bodies, utilized to
generate energy
6. Deamination of amino acids (reductive, hydrolytic, intramolecular, oxidative). Direct
and indirect oxidative deamination. Reductive amination.
Deamination of amino acids is the removal of amino group as free ammonia to prepare
for excretion.
Reductive deamination: conversion of carbonyl group (aldehyde/ketone) to an amine via
intermediate imine.
Hydrolytic deamination: transform cytosine into uracil, adenine into hypoxanthine and
guanine into xanthine and release free ammonia
Oxidative deamination of glutamate: takes place inside mitochondria of hepatocytes
and coupled with oxidation. Glutamate is transported from the cytosol to mitochondria
and undergoes catalyze of glutamate dehydrogenase (using NAD and NADP as
cofactor):
Role: provide NH3 for urea synthesis and a-keto acid for variety reactions.
• Direct oxidative deamination:
- D or L-amino acid oxidase: use flavin (FMN or FAD) as cofactor
• Indirect oxidative deamination: 2 stages:
- 1st: transamination: converts alanine and a-ketoglutarate into pyruvate and glutamate
(catalyzed by alanine aminotransferase)
-
2nd: glutamate under catalyzed of glutamate dehydrogenase turns into a-ketoglutarate
and releasing ammonia. A-ketoglutarate then goes to transamination.
Reductive amination: one of major pathways of ammonia detoxification
- A step in the biosynthesis of many α-amino acids of an α-ketoacid, usually by
a transaminase enzyme. The process is catalyzed by pyridoxamine phosphate.
- The initial step is formation of an imine, but the hydride equivalents are supplied by a
reduced pyridine to give an aldimine, which hydrolyzes to the amine
7. Ttransamination reactions. The role of vitamin B6. The reactions catalyzed by
aspartate aminotransferase (AST) and alanine aminotransferase (ALT). Biologic role.
Diagnostic value.
Transamination: chemical reaction that transform amino acid and alpha keto acid into
new amino acid and keto acid. This process takes place in cytoplasm
NO RED AROW
Role: converts essential amino acids to non-essential amino acids
Mechanism:
- Transfer amino group from aa1 to coenzyme PLP to form pyridoxamine phosphate.
Aa1 is converted to keto acid 2
- Amino group of pyridoxamine phosphate is transferred to keto acid 1 then produce
new aa2 and coenzyme PLP is regenerated
Vitamin B6
Dietary sources: liver, meat, egg yolk, cereal, green vegetables
Daily requirements: 1.4-2.0mg/day
Biochemical role:
- Cofactor enzyme in transamination reaction in synthesis and catabolism of amino
acid process: carrier of amino groups at the active site of aminotransferase and
donate it to keto acid.
- Cofactor for synthesis of neurotransmitter GABA
Functions: cofactors for all transamination and GABA synthesis
Deficiency signs: anemia, homocysteinuria, tuberculosis
Alanine aminotransferase: catalyzes: Alanine + a-ketoglutarate ↔ pyruvate + glutamate
Role: Converts alanine into pyruvate. High level of AAT in blood shows liver damage
Aspartate aminotransferase catalyzes
Role: Converts aspartate into oxaloacetate. High level of AST in blood shows liver damage
Increasing AST- myocardial infarction
Increasing AST, ALT – hepatitis, alcoholic cirrhosis
8. Synthesis of biogenic amines (γ-aminobutyric acid, histamine, serotonin, dopamine).
The role of these biogenic amines. Catabolism of biogenic amines: deamination with
monoamine oxidase (MAO) and methylation reactions.
Synthesis of biogenic amines: decarboxylation of amino acids: reduction of carbon group
in amino acid to yield corresponding amine. Using enzyme decarboxylase and coenzyme
PLP
Synthesis: forming corresponding amine from amino acid
Roles: important hormones, neurotransmitters in human body
Catabolism of biogenic amides:
Monoaminooxidase:
- Cofactor: FAD
- Remove amine group and result ketone or aldehyde and ammonia
- Break down serotonin, melatonin, dopamine, epinephrine, tyramine, tryoamine, etc
Methyl transferase:
- Catalyzes transfer of methyl group
9. Sources of ammonia in tissues. Ammonia toxicity. Transport forms of ammonia in
blood (glutamine, asparagine, alanine). The reactions of their formation. Glucosealanine cycle. The role of the liver, kidney and intestines in the binding and elimination
of ammonia.
Main sources of ammonia in tissues
- Deamination of amino acids
- Deamination of nitrogen bases of nucleotides
- Deamination of biogenic amines
- Product metabolism of bacteria in large intestine
Ammonia toxicity:
- Accumulation of glutamine
- Alteration of neurotransmitter system (GABA, GLU)
- Decrease of energy supplement
- Alkalosis
- Hypoxia
Transport forms of ammonia in the blood:
Glutamine: (major)
- Ammonia is transported from tissue to the liver under form of glutamine (which has
electrolyte balance so it can easily move into the liver)
- Ammonia + glutamate -glutamine synthase-> glutamine
- After get to liver, glutamine undergoes catalyzed of glutaminase or glutamate
dehydrogenase and yield free ammonia to form urea.
Asparagine is a major recipient of glutamine nitrogen and provides a mobile reservoir for
transport to sites of growth
Glucose-alanine cycle:
- Localization: skeletal muscle and liver
- Enzyme: alanine transaminase
- Role: transport a-amino group which is removed from amino acid in the skeletal muscle
to the liver for metabolism.
- Reactions:
+ In muscle: glucose under glycolysis process is converted into pyruvate, Pyruvate
together with accumulate ammonia in muscle transform into alanine.
+ Alanine passes through blood stream goes to the liver
+ In liver: alanine releases ammonia and transformed into pyruvate. Pyruvate under
gluconeogenesis is converted into glucose
10. Urea cycle: localization, enzymes, biologic role. Interaction between urea cycle and
TCA cycle. Diagnostic value of urea concentration in blood and urine. Reference range.
Urea cycle:
- Localization: cytosol and mitochondria in liver
* In mitochondria in liver
+ The first reaction: from carbondioxide and ammonia forming carbamoyl phosphate.
This reaction using 2 ATP
+ The second reaction: Carbamoyl phosphate together with ornithine forming citrulline.
This reaction is catalyzed by ornithine transcarbamoylase.
* In cystosol in liver
+ The third reaction: citrulline and asparagine under catalytic of arginosuccinate
synthetase forming argininosuccinate. This reaction uses 1 ATP. Aspartate arises from
transamination reactin between oxaloacetate and glutamate.
+ The fourth reaction: argininosuccinate is cleaved to arginine and fumarate by
argininosuccinate lyase.
+ The end reaction: is the cleavage of arginine forming urea and ornithine catalyzed by
arginase. Ornithine can go to mitochondria and performs as an intermediate of the
cycle.
- Energy requirement: 3 ATP (2 ATP for 1st reaction, 1 ATP for 3rd reaction)
- Net equation:
2NH3 + CO2 + 3ATP + H2O→Urea + 2ADP + AMP + 2H3PO4 + H4PO7
- Role: Urea contains 2 nitrogen atoms which are derived from ammonia and aspartate.
Urea is well soluble in water and nontoxic substance that can enter the blood and be
excreted with urine. Urea is suitable for detoxification ammonia of the human body
without toxic the blood.
Interaction between urea cycle and TCA cycle: produced fumarate in urea cycle (4th
reaction) can enter mitochondria to participate the citric acid cycle
Diagnosis urea level in blood and urine:
Reference level is 2-6.5 mmol/l
- Increase: enrich protein diet, increase proteolysis, renal failure, dehydration of tissues
- Decrease: low level of protein in diet, disorder absorption in intestine, injury of liver
11. Hyperammoniemia, their causes and consequences. Physiologic concentration of
ammonia in the blood. Causes of ammonia toxicity.
Hyperammonemias: accumulation of ammonia in blood. 3 types
- Primary (hereditary):
- Secondary (acquired)
- Transitory
Causes: lack of enzymes involved in synthesis of urea cycle
- Type I hyperammonemia is due to defect in carbamoyl phosphate synthetase
- Type II hyperammonemia is due to defect in ornithine transcarbamoylase
- Citrullinuria is due to defect in argininosuccinate synthetase
- Hyperargininemia is due to defect in arginase
Clinical manifestations: episodic encephalopathies such as convulsion and ataxia may
occur in children with these diseases.
Principles of treatment:
- Strict dietary control and supplement of essential amino acids (low protein diet)
- Administration of sodium benzoate and sodium phenylacetate. It is beneficial in
reducing level of ammonia serum. These compounds react with glycine and glutamine to
form adducts, which are excreted in the urine
Benzoic acid + Glycine → Hippuric acid
Physiologic concentration of ammonia in the blood: 15-45 micromol/dL
Causes of ammonia toxicity: disorder in detoxification or overproduction of ammonia
- Lack of enzymes for urea cycle, synthesis of biologic amines, reductive amination of
alpha ketoglutarate, formation of ammonia salts
12. The purine and pyrimidine nucleotides: structure, functions in the body. Sources of
nitrogen and carbon atoms in the purine ring. The scheme of synthesis of purine
nucleotides, regulation. Synthesis of deoxyribonucleotides. Enzymes. Role of NADPH
and thioredoxin. Reaction dTMP synthesis. The role of folic acid and tetrahydrofolate.
The cause of megaloblastic anemia in folate deficiency. The mechanism of the
antibacterial activity of sulfanilamides. Inhibitors of timidyl nucleotide synthesis methotrexate, 5-fluorouracil, azidothymidine.
Purine:
- Inosine monophosphate: precursor of AMP and GMP (Adenosine monophosphate
Guanine monophosphate)
Structure: IMP: contain purine ring and ribose 5-phosphate
AMP and GMP: contains purine ring built on PRPP (5-phosphoribosyl 1-pyrophosphate)
and purine base (adenosine, guanine)
(IMP) (IMP: OH, AMP: NH2, GMP: OH and NH2)
Role: components of DNA and RNA
Pyrimidine nucleotides: derivatives of pyrimidine, nucleobases in nucleic acids. They
can form hydrogen bonds with complementary purines: A=T, G-C
- Orotidine 5-monophosphate (OMP)
- Uridine 5-monophosphate (UMP)
- Cytidine 5-monophosphate (CMP)
- Deoxythymidine 5-monophosphate (dTMP)
Sources of nitrogen and carbon atoms for synthesis of purine ring:
+ Glutamine, glycine, aspartate, carbon dioxide, active formiate (from tetrahydrofolate)
Synthesis and regulation of purine nucleotides
Synthesis of deoxyribonucleotides: Reduction of ribose portion of ribonucleoside
diphosphate catalyzed by ribonucleotide reductase.
Ribonucleotide reductase complex of enzymes: (dADP, dGDP, dCDP, dUDP): catalyze
reaction converting ribonucleotide into deoxyribonucleotide which is building block in
DNA replication.
Synthesis of dTMP: dTMP (deoxythymidine 5-monophosphateis) is synthesized from
dUMP in reaction catalyzed by thymidylate synthase
Role of folic acid and tetrahydrofolate:
Folic acid:
- Support DNA synthesis through generation of nucleic acid building blocks
- conversion of homocysteine to the amino acid methionine
Tetrahydrofolate
- Main active metabolite of dietary folate. It is vital as a coenzyme in reactions involving
transfers of single carbon groups.
- Role in nucleic and amino acid synthesis. As nucleic and amino acid synthesis is
affected by a deficiency of tetrahydrofolate, actively dividing and growing cells tend to
be the first affected.
Megaloblastic anemia due to folate deficiency:
- Folic acid plays important role in DNA synthesis by converting homocysteine to aa
methionine. Deficiency in folic acid shows abnormality in RBCs precursor which are
about to divide
- There is no cell division and cytoplasmic accumulation remain unhampered lead to
big sized abnormal cells and too many precursor cells die lead to anemia
Antibacterial properties of sulfonamides
- Stop bacteria from reproducing but don't necessarily kill the bacteria
- Work by interfering with the synthesis of folic acid in bacteria, which is essential for
nucleic acid formation and ultimately DNA and RNA
Inhibitors of thymidyl nucleotide synthesis - methotrexate, 5-fluorouracil,
azidothymidine.
- Methotrexate: inhibits dihydrofolate reductase activity (catalyzes reduction of
dihydrofolic acid to tetrahydrofolic acid). practical apply for cancer treatment
- 5-fluorouracil: inhibits thymidylate synthase (catalyzes convertion of dUMP to dTMP).
practical apply for cancer treatment
- Azidothymidine: Inhibits telomerase which is a region at the end of chromosomes.
Prevention and treatment of HIV/AIDS.
13. Catabolism of purine nucleotides: decomposition of AMP and GMP; reaction of
reutilization of hypoxanthine and guanine, the reaction of formation of uric acid from
hypoxanthine and xanthine, the role of xanthine oxidase. Primary and secondary
hyperuricemia, their causes and consequences: urolithiasis: causes, biochemical aspects
of pathogenesis and treatment; gout: causes, clinical manifestation, biochemical aspects
of pathogenesis and treatment. The mechanism of allopurinol effect in the treatment of
gout. Lesch Nyhan syndrome, the causes, the principles of treatment, prognosis.
Catabolism of purine nucleotides (AMP and GMP):
AMP and GMP catabolism:
- 1st step: dephosphorylation. Nucleotide loses their phosphate group through action of
5-nucleotidase and yield nucleoside (adenosine and guanine)
- 2nd step: catalyzed by nucleoside phosphorylase, yield free nitrogenous base (guanine,
adenine)
- 3rd step: free nitrogenous base under catalyzed of deaminase yield inosine.
- 4th step: inosine transforms into hypoxanthine by nucleoside phosphorylase (only for
adenine)
- 5th step: hypoxanthine is oxidized to xanthine catalyzed by xanthine oxidase (guanine
deaminase converts guanine into xanthine)
- 6th step: xanthine is oxidized to uric acid by xanthine oxidase.
- Uric acid is final product of purine degradation and excreted with urine
Xanthine oxidase: a flavin linked enzyme with atom of molybdenum included in the
structure of its active site. This enzyme also contains iron and sulfur in active site
Reutilization of hypoxanthine and guanines, their roles
Hypoxanthine + PRPP → IMP
Guanine + PRPP → GMP
Role: reduces the need and energy expenditure of biosynthesis by maintaining nucleotide
levels.
Hyperuricemia: excess level of uric acid in blood. The max level of uric acid in blood is
6.8mg/dL
- Primary hyperuricemia: increase uric acid level due to purine
- Secondary hyperuricemia: increase uric acid level due to other factors (diseases or
medications)
Causes:
- Primary hyperuricemia: kidney cant get rid of uric acid in the blood
- Secondary hyperuricemia: chemotherapy, medications, kidney diseases,..
Clinical manifestations: kidney problems, gouty arthritis, uric crystal in joints
Principle of treatments:
- Avoid high purine diet, eat food that are low in purine
- Medication: xanthine oxidase inhibitors (allopurinol)
Urolithiasis: forming stones in kidney, bladder or urethra
- Causes: urinary stasis, metabolic derangement
- Manifestations: abdominal pain, urinary tract infection, blood in urine
- Principle of treatment: keep hydrate for easier excrete stone out, medication (nonsteroid anti-inflammation)
Gout: inflammation of joints. Uric acid crystalized, forming crystal deposits in joints,
tendons.
- Causes: the result of hyperuricemia
- Clinical manifestations: pain, swelling, redness in joints.
- Principle of treatment: less purine diet, keep hydrated, medications (allopurinol)
Allopurinol can inhibit xanthine oxidase, which is enzyme converting hypoxanthine and
xanthine into uric acid. It can help to reduce amount of uric acid in the blood, to treat
gout
Lesh-Nyhan syndrome:
- Causes: genetic disorder results in deficiency of hypoxanthine guanine phosphoriboxyl
transferase
- Clinical manifestation: gout, mental retarded, kidney stone, hyperuricemia, uric acid
crystal in urine, tophi
- Treatments: medication: alloourinol and febuxostat
14. The scheme of pathways of glycine and serine. Interconnection of metabolism of
glycine, serine, methionine and cysteine, vitamins B6, B9 and B12: reaction of
interconversion of serine and glycine, formation of methylene and methyl
tetrahydrofolate, S-adenosyl methionine synthesis from homocysteine, the role of
vitamin B12; S-adenosyl methionine in transmethylation processes for the synthesis of
biologically important substances; reaction of homocysteine production and the
pathway of its transformation into cysteine; role of vitamin B6.
Metabolism of glycine and serine
↓
Oxolate, formylCoA
+ pyruvate, NH3
+ glycine
+ glycerate -> glucose
+ choline
• Glycine: precursor for heme synthesis, ….
- Under oxidative deamination or transamination, glycine is transformed into
glyoxylic acid. This acid undergoes decarboxylation forming formyl FH4
• Serine
- Serine is formed from 3-phosphoglyceric acid (intermediate metabolism of glycolysis)
- Serine is formed from choline through ethanolamine
- Under serine dehydratase, serine is deaminated to pyruvic acid
- Decarboxylation of serine gives ethanolamine
Serine-glycine interconversion catalyzed by serine transhydroxymethylase. This reaction
requires pyridoxal phosphate and tetrahydrofolic derivative (FH4) as coenzymes
Methionine-cysteine interconversion: S-adenyl methionine: - methyl group transfer,
regulate DNA and RNA
Vit B6
- Needed for
cystothionine synthase
and cystothionase as
coenzyme
- Transamination
decarboxylation
sphingolipid synthesis
- Precursor for pyridoxal
phosphate (PLP) and
pyridoxamine phosphate
(PAMP)
Vit B9: Folic acid
- Required for Gly-serine
interconversion as
coenzyme (conversion of
methylene and methyl
tetrahydrofolate)
- Imp for rapid dividiy all
- Synthesis of puridine,
pyrimidine
Coenzyme form:
tetraphydrofolic acid
(THFA)
Avitaminosis:
Avitaminosis:
- Pellagra – like dermatitis - Megablastis anemia
(not sensitive)
- Neural tube detect
Vit B12: cobalamin
- Normal maturation RBC
- Transfer methyl group
(methyl transferase in
met-cys interconversion)
Coenzyme form:
- Methylcobalamine
(acetyl B12)
- Deoxyadenosyl
cobalmime
Avitaminosis:
- Megaloblastic anemia
- Tubercullosis homocysteinuria
S-adenyl methionine synthesis from homocysteine
In methylation, SAM acts as methyl donor to help produce and regulate hormones and
maintain cell membrane
15. Metabolism of phenylalanine and tyrosine. Anabolic and catabolic pathways of
tyrosine transformations. The reaction converting phenylalanine to tyrosine.
Characteristics of disease phenylketonuria type 1 (classic) and phenylketonuria type 2
(variant): defective enzymes, biochemical aspects of pathogenesis, typical clinical
manifestations, principles of treatment.
Phenylalanine: Standard aa, essential aa that dont synthesis in body (tissue) only 1 way
provided by food protein, glucogenic aa
Main pathway utilization:
- protein synthesis and peptides (most aa use this)
- maybe use for synthesis non essential aa Tyrosine (using phenyl alanine hydrolase)
- decompostion resulted from yield of product catabolism such as amino water.
Tyrosine: Standard, nonessential aa, glucogenic aa.
- Tyrosine can be synthesized from Phe (essential aa)
Fumarate
- Involves in CAC, intermediate CAC: use for synthesis acetylacetate (for gluconeogenesis)
- undergo complete decomposition accompanied by release energy use for synthesis ATP
Acetoacetate
- Undergoes conjugation with CoA: transformed into acetoacetyl CoA under catalyze of
3ketoacyl CoA transferase
Tyrosine maybe used for
- Synthesis of protein and oligopeptides
- Synthesis of catecholamine (dopamine, NE, epinephrine: biological substances used as
hormones, neurotransmitters)
- Synthesis thyroid hormone
- Synthesis pigment in skin, eyes area (melanin)
Phenylketonuria:
- Type 1: caused by genetic defect of phenylalanine hydrolase enzyme
- Type 2: caused by defect in dihydrobiopterin reductase which is needed for regeneration
of the cofactor for hydroxylation of phenylalanine
- Mechanism: phenylalanine from diet cannot be converted into utilized form in human
=> accumulation in blood and human tissues => increase toxic level
- Symptoms: musty odor in breath, skin, urine; neurological problems: seizures’
hyperactivity, delayed development, fair skin and blue eye (Phe cant transform into
melanine)
- Biochemical diagnosis: urine and blood test for Phe level, cofactor deficiency testing
(BH4), PAH detect in hepatic and renal tissues
- Principles of treatment: lifetime diet with limited protein intake, and addition of
supplement for essential protein but wo Phe
16. The reactions of conversion of tyrosine to dihydroxyphenylalanine, dopamine,
norepinephrine and epinephrine. Disorders of tyrosine metabolism - albinism and
parkinsonism. Biochemical aspects of pathogenesis and treatment.
Catechoamine: epinephrine,
norepinephrine, dopamine
Role:
- Regulate carbohydrate and lipid
metabolism
- Stimulate degradation of triacylglycerol
and glycogen
- Cause increasing blood pressure
- Dop and NE serve as neurotransmitter in
brain and ANS
Albinism:
- Causes: defective enzyme: tyrosine 3 monooxygenase, defective process: melanin
synthesis from tyrosine
- Symptom: lack of pigmentation skin, hair, eyes
- Biochemical aspects of pathogenesis: melanin is color pigment. The deficiency in
forming melanin causes the lack of pigmentation throughout the whole body (eyes, skin,
hair)
- Principles of treatments: symptom management: prevent sun damage
Parkinsonism:
- Causes: deficiency of dopamine from disorder of nerve cells
- Biochemical aspects: Dopamine inhibits human movement. Lack of dopamine leads to
uncontrol movement
- Symptoms: shaking, stiffness, difficulty walking, balance, etc
- Principles of treatment: supplement of dopamine
17. Metabolism of arginine. Arginine in the synthesis of urea, creatine, nitric oxide (NO).
Reactions of polyamines synthesis (spermine and spermidine). The structure of creatine
and creatine phosphate, the reaction of their synthesis, localization of the process. The
biological role of creatine phosphate.
Histone role:
- Package DNA into structural unit => nucleosomes
- Gene regulation
Creatine and Creatine phosphate
Localization: muscle (creatine phosphate -> creatinine) and they are transported through
blood to kidney for elimination
Role:
Creatine
- Chemical waste product (by muscles)
- Creatine level in blood reflect amount of muscle of a person and the health of kidney
- Increase level of creatine = kidneys do not function well (kidney disease = high level of
creatine or urea)
Creatine phosphate
- Form of energy store
- Provide phosphate group rapidly in disease
MODULE VI. HORMONES. HORMONAL REGULATION OF METABOLISM
1. Hormones. Hierarchy of regulatory systems. Classification of hormones according to
chemical structure. Cell signaling. Plasma membrane receptors and intracellular binding
proteins. Intracellular second messengers.
Hormones and hierarchy of regulatory system
Hormones are biologically active substances synthesized in endocrine glands. They are
secreted to the blood stream and have a regulatory effect on the metabolism and function
of visceral organs
Hierarchy of regulatory systems
Hipothalamus
Anterior pituitary gland
Posterior pituitary gland
Endocrine glands (pancreas, thyroid, parathyroid, adrenal cortex, pineal gland)
Target cells in visceral organs
Classification of hormones according to chemical structure:
- Peptide hormones (include peptide, simple and complex proteins): maybe in inactive
form, in prohormone form (insulin, glucagon, etc). They are accumulated in gland tissue
and secreted in active form. Formation of active form associated with proteolysis which
means reducing molecular mass.
- Amino derivatives (epinephrine: from tyrosine, thyroid hormone,etc)
- Steroid hormones (sex hormones and adrenal cortex hormones): derived from
cholesterol
2. The adenylyl cyclase mechanism of hormonal action: hormones, second messenger,
enzymes and processes regulated by this mechanism. Reaction of cAMP synthesis and
breakdown. Activation of protein kinase A. The role of activating and inhibitory isoform
of α subunit of G protein. The transcription factor CREB. Phosphatidylinositol signalling:
hormones, second messengers, enzymes and processes regulated by this mechanism.
The reaction of formation of inositol triphosphate (IP3) and diacylglycerol (DAG).
Sources of calcium ions.
Adenylate cyclase mechanism:
- Hormones: epinephrine (adrenal medulla)
- Second messenger: cAMP
- Enzyme: adenylate cyclase
- Processes:
+ Hormone binds to adrenergic receptor forming hormone-receptor complex
+ The hormone-receptor complex is accompanied with change is structure of G protein.
G protein breaks its 3 subunits into monomer (alpha subunit) and dimer (beta and gamma
subunits)
+ alpha subunit binds to molecule GTP and they act as activator of adenylate cyclase
enzyme.
+ Adenylate cyclase enzyme catalyzes ATP hydrolysis reaction with formation of cAMP
ATP + H2O => cAMP + H4P2O7
+ cAMP exhibits properties of an allosteric activator of enzyme protein kinase A
(catalyzes phosphorylation reaction of intracellular protein using ATP)
G protein: is a trimer protein which located on the inner surface of membrane
(cytoplasm). G protein consists of 3 subunits (alpha, beta and gamma: 3 diff polypeptide
chains)
- When GDP binds to the alpha subunit, it remains binding to beta and gamma subunits
- When GTP binds to the alpha subunit, the alpha subunit dissociates from the betagamma complex and is then free to interact with other effector molecules
Role of activator and inhibitor isoform of alpha subunit of G protein:
- Formation of hormone-receptor complex changes structure of G protein and makes it
split into alpha and beta, gamma subunit.
Metabolism of cAMP
- cAMP is synthesized by ATP hydrolysis reaction under catalization of adenylate cyclase:
ATP + H2O => cAMP + H4P2O7. cAMP is allosteric activator of enzyme protein kinase A.
- In phosphodiesterase reaction, cAMP is converted into noncyclic AMP
cAMP + H2O => AMP
AMP has no property of second messenger of hormone
Mechanism of protein kinase A activation
- Protein kinase A catalyze the phosphorylation reaction using ATP. It consists of 4
subunits: 2 regulatory and 2 catalytic.
- When 4 cAMP bind to binding sites in regulatory subunit, the enzyme is dissociated
into 4 separated subunits.
- Catalytic subunits now can perform catalytic properties of this enzyme
The transcription factor CREB: activated by cAMP, is a cellular transcription factor. It
binds to cAMP and works in nucleus and thereby regulate the increasing or decreasing
transcriptions of genes
Phospholipase C mechanism:
- Hormones: vasopressin, gastrin, thyroliberin, gonodoliberi
- Second messenger: inositol triphosphate (IP3), diacylglycerol (DAG), and calcium
- Increase of Ca level in cytoplasm mechanism:
+ Forming hormone-receptor complex is accompanied by the change in subunit of G
protein resulting in activation of effector protein phospholipase C in inner surface of
cell membrane
+ Phospholipase C catalyzes reaction of phosphatidyl inositol (IP3) hydrolysis
IP 4,5 biphosphate => 1,2 diacylglycerol + inositol 1,4,5 triphosphate
+ Inositol triphosphate interacts with receptor in endoplasmic reticulum of the cell,
forming a complex and promotes calcium releasing from ER to cytoplasm.
+ Calcium acts as second messenger, forms complex with calmodulin protein in
cytoplasm. This complex activates numbers of enzyme: phospholipase A2,
phosphodiesterase, pyruvate kinase
3. Receptors associated with tyrosine kinase activity: enzymatic cascade that is
associated with the activation of Ras-protein, its scheme, the sequence of events, the
main participants, the role for cell metabolism. Cytosolic mechanism of hormonal
signals: stages, hormones, features of intracellular receptors.
Receptor associated with tyrosine kinase
Example: insulin, oxytoxin
Receptor has 4 subunit, 2 alpha and 2 beta type.
- Alpha subunit located out of cell membrane: responsible for recognition and binding of
hormones
- Beta subunit is transmembrane: exhibit properties of tyrosine kinase enzyme catalyzing
the reaction of tyrosine residues phosphorylation by ATP
Intermediate substance: Ras-protein: G protein: contains 3 subunits: alpha, beta and
gamma.
Ras protein is activated, it stimulates the tyrosine kinase, help change DNA => RNA =>
changes in enzyme (forming new enzyme)
Cytosolic (intracellular) mechanism
Mechanism:
- Hormone penetrates through the hydrophobic layer of cell membrane and enters the
cytoplasm.
- Hormone binds to receptor in cytoplasm and forms the complexes hormone-receptor.
- This complex moves to nucleus and binds to specific sites of chromatin (promoter sites
on DNA) and results the corresponding gene synthesis is stimulated.
- Then, level production of certain protein (include enzymes) increases.
Eg: Steroid hormones regulate synthesis of gonadotropin hormones, integrins, growth
factors, and inflammatory response proteins.
4. Hypothalamic-pituitary-adrenocortical system: biological role, components,
regulation. Glucocorticoids: regulation of synthesis and secretion, the main stages of the
synthesis, mechanism of action and target organs, the effect on metabolism - regulated
processes. Hypo- and hyperproduction of glucocorticoids - metabolic disorders,
biochemical aspects, principles of treatment.
Hypothalamic-pituitary-adrenocortical system:
-Brain: stressful signal: pain, hypoglycemia, hypoxia
(+)
-Paraventricular nucleus of hypothalamus: corticotropin-releasing hormone (CRH)
(+)
-Anterior pituitary: andrenocorticotropic hormone (ACTH)
(+)
-Adrenal cortex: cortisol synthesis => pregnenolone (under cholesterol transformation)
=> transformation into glucocorticoids
High content of lower substances can give negative feedback to higher level hormones.
Glucocorticoids
Representatives: cortisol, corticosterone
Structure: steroidal structure
Basic synthesis stages:
-ACTH reaches specific receptor on plasma membrane of cells situated in zona
fasciculata of adrenal cortex through bloodstream and brings to transformation of
cholesterol into pregnenolone
-Pregnenolone simultaneously transformed into cortisol and corticosterone
Regulation of synthesis and secretion:
-Under strict control of cascade of neural endocrine signals linked with cerebro-corticalhypothalamic-pituitary-adrenocortical system
-High level of cortisol in the blood suppresses CRH and ACTH secretion
-Low level of cortisol in blood stimulates CRH and ACTH secretion
Mechanism of effects on the target cells
-Binding to cytosolic receptors which then act to modulate gene transcription in target
tissues.
Metabolic effects
-Cortisol inhibits uptake and utilization of glucose resulting in increasing of blood glucose
level. Further, it increases the breakdown of skeletal muscle protein and adipose tissue
TAG to provide energy and substrates for gluconeogenesis
-Increasing of protein metabolism rate leads to increasing urinary nitrogen excretion and
induction of urea cycle enzymes.
Using as medicines
Glucocorticoids have immune suppressive and anti-inflammatory effects. They are used
in form of related drugs such as prednisone for treatment of inflammation and allergy.
Hypoproduction of glucocorticoid
Metabolic disorders: Addison’s disease (the immune system produces abnormal
antibodies that attack cells of the adrenal cortex), untreated infections of adrenal cortex,
decrease output of ACTH due to pituitary tumor
Clinical manifestation: hyperpigmentation of the skin, fatigue, anorexia, orthostasis,
nausea, muscle and joint pain, and salt craving.
Principles of diagnostic:
- Metabolic test: test cortisol level and determine whether the adrenal insufficiency is
primary or secondary
- Confirming an elevated ACTH level and an inability to stimulate cortisol levels with a
cosyntropin stimulation test
Treatment: treated with replacement hormones in pill form
Hyperproduction of glucocorticoid
Causes: pituitary tumor or adrenal cortex tumor that leads to excessive production of
ASTH
Metabolic disorders: Cushing syndrome, hypertension, diabetes mellitus, obesity and
anorexia, osteoporosis and immune disorders, several psychiatric disorders, Alzheimer
disease
Clinical manifestation: stretch marks in the skin, skin becomes progressively thinner,
moon face which fat deposits give the face a rounded appearance, thin hand
Principles of diagnostic:
- Urinary and blood test to measure hormone level
- Saliva test: Cortisol levels normally rise and fall throughout the day. In people without
Cushing syndrome, levels of cortisol drop significantly in the evening
Treatment: depends on its cause and may include surgery to remove a tumor or
medications to suppress activity of the adrenal glands
5. Hypothalamic-pituitary-thyroid system, the biological role, components, regulation.
Thyroid- stimulating hormone: regulation of synthesis and secretion, structure,
mechanism of action and target organs, biological effects.
Hypothalamic-pituitary-thyroid system
Biological role: maintain normal, circulating levels of thyroid hormone that is essential for
the biological function of all tissues, including brain development; regulation of
cardiovascular, bone, and liver function; food intake; and energy expenditure.
Component and regulation:
- Hypothalamus: Thyrotropin-releasing hormone (TRH)
(+)
- Anterior pituitary: Thyroid stimulating hormone (TSH)
(+)
(-)
- Thyroid: T3 and T4 High level of T3 and T4
Thyroid stimulating hormone
TSH: peptide hormone controls the thyroid hormones synthesis and secretion
Structure: It consists of two chains: an alpha chain and a beta chain. It has a molecular
mass of approximately 28,000 Da
Regulation of synthesis and secretion
- Its synthesized in anterior pituitary gland and secreted by pituitary gland
- (-) level of T3 in the blood is over critical level
- (+) Thyrotropin-releasing hormone in neuron of hypothalamus
Mechanism of effects on target organs
- The hypothalamic-pituitary axis regulates TSH release. Specifically, neurons in the
hypothalamus release TRH, or thyroid-releasing hormone, which stimulates thyrotrophs
of the anterior pituitary to secrete TSH.
- TSH, in turn, stimulates thyroid follicular cells to release thyroid hormones in the form
of T3 or T4.
- T3 and T4 can give negative feedback to anterior pituitary gland with high level in blood
Metabolic effects
- Increase the secretion of thyroid hormones
- Prolong the growth of thyroid glandular tissue itself (hypertrophy or hyperplasia)
6. Thyroid hormones: chemical structure, regulation of synthesis and secretion, the
main stages of the synthesis, mechanism of action and target organs effect on
metabolism: processes, regulated by thyroid hormones. Hypo-and hyperthyroidism metabolic disorders, biochemical aspects. Principles of treatment.
Thyroid hormones
Representatives: 3,5,3’-triidotrhyronine (T3) and 3,5,3,5’-tetraiodothyronine
Structures: iodinated dityrosine derivatives
Transported in blood
- T3 and T4 are hydrophobic so they bound to glycoprotein known as thyroxin-binding
globulin and are disseminated throughout the body in this complex.
Basic synthesis stages
- Precursor protein thyroglobulin which localized in thyroid follicles. It contains more than
100 tyrosine residues.
- Thyroglobulin is exocytosed through apical membrane into the closed lumen of the
thyroid follicles and store here.
- Na/K ATPase pumps concentrate iodide (I-) in thyroid cell and transports it to lumen.
- Iodide is oxidized to I+ by thyroperoxidase and added to thyroglobulin, forming
monoiodotyrosyl (MIT) and diiodotyrosyl (DIT)
- Combining of MIT and DIT forming T3 and T4 (but still in complex)
- Mature thyroglobulin is taken up by thyrocytes vesicle and fuse with lysosome.
Lysosomal protases degrades thyroglobulin and releasing T3, T4 and free amino acid into
the circulation.
Regulation of synthesis and secretion
- (+) TSH
Mechanism of effects on target organs
T3 and T4 bind to specific nuclear receptor sites. The nuclear action of T3 results in
organ-specific increases and decreases of specific mRNAs, leading to alteration in the
level of the corresponding proteins. In addition to the well established nuclear action of
T3, effects of thyroid hormone on other sites including cell membranes and
mitochondria have been documented.
Metabolic effects
- Increasing oxygen consumption in most of the body tissues
- Promote protein synthesis by enhancing transcriptional level
- Promote intestinal absorption and utilization of glucose
- Increase blood glucose level by activation of glycogenolysis and gluconeogenesis
- Stimulate of lipid turn over
- Regulation of water and electrolyte metabolism
Using as medicines: using for hypothyroidism
Hypo- and hyperthyroidism
Hypothyroidism: condition with decreasing level of thyroid hormones
- Metabolic disorders: swelling mucosa of skin, subcutaneous tissue, drowsiness,
weakness, bradycardia. Congenitial hypothyroidism results from CNS affection and
growth retardation with irreversible mental retarded
- Biochemical aspects: reduced basal metabolism, decrease in oxidation reaction rate,
decrease heat production.
- Causes
+ Primary: caused by gland tissue damage (tuberculosis, autoimmune thyroiditis,
resection of thyroid gland)
+ Secondary: caused by inhibition of TSH secretion by anterior pituitary (brain damage,
pituitary adenoma)
- Principle of diagnosis: measure of thyroid hormone content in blood.
+ For secondary hypothyroidism: measure blood content of T3 and T4 after injection of
throtropin.
- Treatments: taking daily hormone replacement tablets called levothyroxine
Hyperthyroidism: increasing production of thyroid hormones and their elevated
concentration in blood
- Metabolic disorders: nervousness, tremor, insomnia, hyperthermia, palpitations,
weight loss,…
- Biochemical: intensification of catabolic processes in organism and lead to negative
nitrogen balance
- Causes: due to formation of immunoglobulin in thyroid gland. Ig bind with TSH
receptor and stimulates production of thyroid hormones and lead to high concentration
of T3 and T4 in blood.
- Principle of diagnosis: measure of thyroid hormone content in blood
- Treatment: can be treated with antithyroid medications (methimazole and
propylthiouracil), radioactive iodine ablation of the thyroid gland, or surgical
thyroidectomy
7. Hormonal regulation of absorptive and post absorptive period. Glucagon: biological
importance, regulation of synthesis and secretion, mechanism of action, target organs,
the effect on metabolism - regulated enzymes and processes.
8. Epinephrine: biological role, chemical structure, reactions of synthesis, regulation of
secretion, adrenergic receptors, their distribution, the mechanism of action, depending
on the receptor, target organs, effects on metabolism depending on receptor: regulated
enzymes and processes. Hypo- and hyperproduction of epinephrine - metabolic
disorders, biochemical aspects. Principles of treatment.
Structure: catecholamine. Precursor: tyrosine.
Synthesis: catecholamine is synthesized in the cells of adrenal medulla (chromaffin cells)
from precursor tyrosine.
Tyrosine -tyrosine hydrolase-> L-dopa -aromatic L amino acid decarboxylase-> dopamine
-dopamine b hydrolase -> norepinephrine -phenylethanolamine N methyl transferase ->
epinephrine.
Regulation of synthesis and secretion:
- (+) pain, hypoxia, exercise, blood loss, hypoglycemia: stimulate to adrenergic nuclei in
hypothalamus => influence the sympathetic on chromaffin tissue of adrenal glands and
release of epinephrine into the blood.
- (-) When the stressful situation ends, the nerve impulses to the adrenal glands are
lowered, meaning that the adrenal glands stop producing epinephrine
Adrenergic receptors
There are two types of adrenergic receptors a and b. Epinephrine is able to interact with
both type of receptors.
- When it forms complex with a receptor, it will decrease the level of cAMP
- When it forms complex with b receptor, it will increase the level of cAMP
Mechanism effects on target organs: hormone that binds to membrane receptors
- Target organs: liver, adipose tissue, skeletal muscle
- When epinephrine-receptor complex forms, it activates the adenyl cyclase which
catalyzes the reaction forming cAMP.
- cAMP activates protein kinase A => activates series of different enzymes (glycogen
synthase, pyruvate kinase, pyruvate dehydrogenase complex, hormone sensitive lipase,
fructose 1,6 biphosphate, phosphofructokinase)
Metabolic effects:
- Epinephrine mediates effects of sympathetic nervous system on the tissues of visceral
organs blood vessels, heart, lungs, gastrointestinal tract and others. Stimulation of
epinephrine results in elevation of blood pressure, increase of cardiac output,
intensification of intestinal motility, dilation of bronchi, etc. All these effects are in
improving the blood supply of the tissues and providing them with substrates for
oxidation
Medicines:
As a medication, it is used to treat a number of conditions, including anaphylaxis,
cardiac arrest, asthma, and superficial bleeding.
Hypoproduction of epinephrine: it is very rare and patients cannot react properly to
stressful situations
Causes: Tyrosine Hydroxylase Deficiency is caused by mutations of the tyrosine
hydroxylase (TH) gene.
Manifestation:
- clumsy manner of walking (abnormal gait)
- dystonia (a group of muscle disorders generally characterized by involuntary muscle
contractions that force the body into abnormal, sometimes painful, movements and
positions (postures))
Metabolic disorder:
- Mutation of the TH gene results in deficient levels of tyrosine hydroxylase, which, in
turn, results in a deficiency of dopamine, norepinephrine, and epinephrine.
Principles of treatment:
- medications to restore normal dopamine levels in the brain
- The drug selegiline has been used to treat individuals with tyrosine hydroxylase
deficiency. Selegiline is a monoamine oxidase type B (MAO-B) inhibitor and slows down
the breakdown (catabolism) of dopamine in the body
Diagnostic: blood test, urine test
Hyperproduction of epinephrine: Pheochromocytomas are tumors that develop in the
adrenal medulla.
Causes: tumors of adrenal medulla, stress
Manifestion: Excess of catecholamines causes high blood pressure. Pheochromocytoma
patient with hypertension. However, if left untreated, it can have serious consequences,
including heart attack, stroke, and other life-threatening conditions.
Metabolic disorder: Epinephrine mediates effects of sympathetic nervous system on the
tissues of visceral organs blood vessels, heart, lungs, gastrointestinal tract and others.
Hyperproduction of epinephrine can make more blood supply to tissues and promote
oxidation
Principle of treatment: remove adrenal tumors surgically
Diagnostic: blood test, urine test, imaging (radiology) tests of the adrenal glands,
9. Insulin: the biological role, the main stages of the synthesis, regulation of secretion,
mechanism of action of insulin, the molecular effects of insulin - the metabolic and
mitogenic pathway.
Insulin: simple hormone, consist of 2 polypeptide chains a and b (oligopolypeptide) with
low molecular weight (only 51 aa). The connections between 2 chains are designated by
link between amino acid Cystein.
Biological role: decrease level of glucose in the blood by stimulating glucose absorption
of cell for different pathway: glycogenesis, PPP, glycolysis
Synthesis:
- Synthesis from beta cell of Langerhans in pancreas. Precursor of insulin is proinsulin.
- Proinsulin consists of 1 polypeptide chain. Its linked between different part of chain by
Cys-Cys connection. Proinsulin under restrict proteolysis (catalyzed by trypsin-like
protease), decrease molecular mass, breaks into insulin and C-peptide.
- Insulin binds with zinc cations and accumulates in special granules.
- Under the influence of certain stimuli, insulin is segregated from pancreas to the
bloodstream and transported to target organs and binds to glycoprotein receptors.
Regulation of secretion:
- Hyperglycemia: Glucose involves in glycolysis (in all tissue of human cell body): bring to
increase synthesis of ATP in b-cell => increase rate of ATP and ADP in b cell => brings to
depolarization of cell membrane => open of voltage dependent Ca channel => Ca influx
=> increase of Ca level inside the cell => activation of phospholipase C => release of Ca
from endoplasmic reticulum. => more increase Ca inside cell => Mobilization of insulin
granule => Insulin together w phospholipase C pass cross membrane get to blood
stream
- Parasympathetic system: increase synthesis of insulin
- Activation a2-adrenergic receptor (NE) brings to inhibition of insulin.
Mechanism of action: insulin binds to receptor and stimulates tyrosine kinase
phosphorylation different cascade
- Metabolic pathway: lipid, glycogen, protein synthesis; cell survival and proliferation;
glucose transport; receptor endocytosis
- Mitogenic pathway: gene expression; transcription factor activation; growth and
differenciation; actin organization
10. Insulin. Very fast, fast, slow, very slow effects. The enzymatic cascade that is
associated with the activation of Ras-protein, its scheme, the sequence of events, the
main participants, role for cell metabolism, enzymatic cascade that is associated with
the activation of phosphoinositol-3- kinase and protein kinase B (AKT), its scheme, the
sequence of events, the main participants, role for the cell metabolism. Glucose
transporters, their types and tissue localization.
Insulin
- Very fast effect: increase glucose and ions transport into the cells
- Fast effect: increase glycogen, fatty acids, glycerol, and protein synthesis
- Slow effect: increase enzyme synthesis that regulate anabolic processes
- Very slow effect: increase cell division
Insulin binding activates insulin receptor --> phosphorylation of insulin receptor
substrates --> IRS protein are recruited to the receptor phosphorylated on tyrosine
residues --> binding sites for molecules with PI3K --> activated PI3K generates
phosphatidylinositol (3,4,5)-triphosphate (PIP3) from phosphatidylinositol 4,5biophosphate (PIP2) --> increasing level of PIP3 activates cascade of different protein
kinase, especially protein kinase B (PKB)
- Enzymatic cascade that is associated with the activation of Ras-protein MITOGENIC
PATHWAY: SH2 and SH3 domains
+ SHC protein are able to bind growth factor receptor-bound protein 2 (GRB2) after
tyrosine phosphorylation => activation of Ras + beginning of a phosphorylation cascade
=> activation of MAPKs (protein used in regulation of cell growth, division, apoptosis and
able to phosphorylate nuclear proteins and several protein kinases that interact with
transcription factors => effect gene expression)
Enzymatic cascade that is associated with the activation of phosphoinositol-3- kinase
and protein kinase B (AKT) METABOLIC PATHWAY
- PKB phosphorylates AS160 which is inhibitor of GLUT4 translocation and brings to
inactivate of AS160. Then GLUT4 translocation occurs, glucose can get into the cell for
glycolysis
- Also, PKB phosphorylates GSK3 and then GSK3 phosphorylates GS (glycogen synthase)
which promote the glycogenesis process.
Role of glucose transporters: Glucose transporters accomplish the movement of glucose
from the extracellular space (deriving from the bloodstream) into cells. The reduction of
glucose in the blood results from the action of insulin.
GLUT1
Placenta, muscle, adipose tissue, brain, endothelium
GLUT2
Pancreatic b-cells, liver, small intestine, renal proximal tube
GLUT3
Neural, small intestine
GLUT4
Muscle, heart, adipose tissue
GLUT5
Small intestine, brain, muscle and adipose tissue
GLUT6
Ubiquitous
11. Insulin. Effect on metabolism of carbohydrates, lipids and proteins.
Insulin belongs to peptide hormone. It is simple hormone, consist of 2 polypeptide
chains a and b (oligopolypeptide) with low molecular weight (only 51 aa). The
connections between 2 chains are designated by link between amino acid Cystein.
Insulin is secreted by b cell of Langerhans in pancreas in precursor proinsulin form.
Insulin influences the glucose intake of cell for glycogenesis, glycolysis, PPP, inhibits
adipose tissue lipolysis and muscle proteolysis
12. Diabetes mellitus type 1 and 2. Causes of absolute and relative insulin deficiency.
Similarities and differences in metabolic disorders in types 1 and type 2 diabetes.
Biochemical mechanisms of clinical manifestations. Causes of insulin resistance.
Biochemical mechanisms of diabetes complications.
Type1 or insulin-dependent diabetes mellitus: is the result of a frank deficiency of
insulin.
Type2 or non-insulin-dependent diabetes mellitus : begin as a syndrome of insulin
resistance
Compare between 2 types of diabetes
Diabetes type 1
Diabetes type 2
Causes
Pancreas is unable to produce
Body does not respond well to
enough insulin
insulin
Age
Normally develop at young age
Normally develop at older age
Prevention Cannot be prevented
Can be prevented with lifestyle
Treatment
Require insulin therapy
Can be managed with lifestyle
Symptoms
Frequent urination increase thirst, extreme hunger, weight loss,
fatigue, blurr vision
Complications Cardiovascular disease, risk of heart attack and stroke, kidney disease
and failure, problem with wound healing, vision loss
Biochemical mechanisms of manifestation:
- High level of glucose in the blood increases blood osmolarity -> blood pressure which
stimulates thirst center. Also, elevated glucose level require urination to get rid of them
from the body
- Glucose is primary fuel for body activities. Unable uptake glucose leads to extreme
hunger and weightloss
Causes of insulin resistance
- Bad habits (obesity, smoking, inactive lifestyle, etc) can lead to disable in insulin
respond in tissue --> body can’t uptake glucose as energy source. To make up for it,
pancreas making more insulin. Overtime, blood glucose increases.
Biochemical mechanism of complications:
- Insulin resistance suppress lipid metabolism --> increase plasma TAG, LDL level --> fatty
foam cells
- Overworking to excrete excess glucose in blood --> kidney disease
- High glucose level in the blood and no energy to synthesis protein for blood clotting -->
problem with wound healing
- High glucose level causes the lenses to swell with fluid --> blurry vision
13. The biochemical diagnostics of diabetes: glucose tolerance test, the concentration of
glycosylated hemoglobin (HbA1c) and C-peptide.
Glucose tolerance test: is a medical test in which glucose is given and blood samples taken
afterward to determine how quickly it is cleared from the blood
- The test is used for test for diabetes, insulin resistance, impaired beta cell function
Concentration of glycosylated hemoglobin (HbA1c): it is uses HbA1c of 48mmol/mol;
(6.5%) to diagnosis diabetes
- The purpose of this method: Glycated hemoglobin (HbA1c) reflect the cumulative
glucose exposure of erythrocytes over a preceding time fram propotional to erythrocyte
survival. HbA1c is an aerial function of the glucose-time curve, an educationally useful
concept to aid teach and clinical judgment
- In patients without symptoms of diabetes the laboratory venous HbA1c should be
repeated. If the second sample is <48mmol/mol (6.5%) the person should be treated as
at high risk of diabetes and the test should be repeated in 6 months or sooner if symptoms
develop.
Concentration of C-peptide: is a useful and widely used method of assessing pancreatic
beta cell function
- The degradation rate of C-peptide in the body is slower than that of insulin => affords
a more stable test window of fluctataing beta cell response
- In healthy individuals the plasma concentration of C-peptide in the fasting state is 0.30.6 mol/l, with a postprandial increase to 1-3nmol/l
- In insulin-treated patients with diabetes, measurement of C-peptides also avoids the
pitfall of cross-reaction of assay between exogenous and endogenous insulin
14. Changes in metabolism of carbohydrates and lipids in fasting and in stress. Stages of
fasting.
Prolonged fasting decreases blood glucose level. Stress activates sympathetic nervous
system with stimulation of epinephrine and cortisol. Long term of these hormones
reduces insulin sensitivity
Body uses secondary source – fatty acids as fuel, forming ketone bodies. Ketone bodies
are acidic, it decreases blood pH. Low pH can lead to dysfunction of proteins in the body
(primary is RBCs)
Stages of fasting
- Stage 1 (8-12h): stable blood sugar
- Stage 2 (12-18h) fat burning --> ketone bodies
- Stage 3 (24h) autophagy
- Stage 4 (36-48h) increase growth hormone stimulating muscle repair
- Stage 5 (72h) trigger new stem cell formation
15. The growth hormone: regulation of the synthesis and secretion, structure, the target
organs, mechanism of action, the role of somatomedins, effect on metabolism regulated processes, hypo- and hyper production - metabolic disorders, biochemical
aspects. Principles of treatment.
Growth hormone
Synthesis and secreted by somatotroph cell in anterior pituitary gland
Hypothalamus
↓(+) GH releasing factor
Anterior pituitary gland
Somatomedin (-) ↓ (+) GH
Insulin-like hormone (liver); increase gluconeogenesis and cell uptake aa, lipolytic effect
Structure: GH composed of a single polypeptide chain with 200 aa residues. There are
several disulfide bonds in polypeptide chain
Target organs: all tissues
Mechanism of action
- Direct: growth hormone binding its receptor on target cells. It acts on fat cells
(adipocytes) and stimulates them to break down triglyceride and supresses their ability
to take up and accumulate circulating lipids.
- Indirect: mediated primarily by a insulin-like growth factor-I (IGF-I), a hormone that is
secreted from the liver and other tissues in response to growth hormone
Hypoproduction
- Metabolic disorder: accumulation of lipids in adipose tissue
- Biochemical aspects: disorder in growing
- Principle of treatment: hormone supplement
Hyperproduction
- Metabolic disorder: inhibit insulin action
- Biochemical aspects: leads to overgrowth
- Principle of treatment: hormone supplement to inhibit GH secretion
Somatomedins: (insulin-like growth factors) group of protein produced predominantly
by the liver when growth hormone act on target tissue
Role:
- Stimulates growth and increases muscle and bone mass, as well as normalising blood
chemistry
- Inhibit growth hormone release and stimulate secretion of somatostatin from
hypothalamus
- In metabolism, somatomedins are competitive inhibitor of insulin and proinsulin
Regulation: autocrine/paracrine
16. Vitamin A: sources, structure, active forms, biochemical functions, clinical signs of
hypo- and hypervitaminosis. Retinoic acid, its receptors, the role in cell differentiation.
Sources: B- carotene (consists of 2 molecules of retinal linked at their aldehyde ends)
(provitamin A)
Active forms: retinal, retinoic acid
Biochemical functions: Takes part in process photoreception. (opsin – covalently linked
with aldehyle form of vitA – 11 cis retinal
Vit A is stored in liver
Hypovitaminosis: Night blindness, follicular hyperkeratinosis, cancer, deterioration of
the eye tissue, xerophthalmia
Hypervitaminosis: blurry vision, bone pain, nausea and vomiting, sensitivity to sunlight,
dry skin
Retinoic acid: a metabolite of vitamin A (all trans retinol)
- Receptor: 3 types: alpha, beta and gamma. They are nuclear receptor which can act as
transcription factor, can be activated by all trans retinoic acid and 9 cis retinoic acid.
The binding to those receptors promote transcription of gene into mRNA and then
produce protein.
- Role in cell differentiation: via retinoic acid receptors, vitamin A can regulate the cell
differentiation and cell growth
MODULE VII. ROLE OF LIVER IN HOMEOSTASIS. BIOCHEMISTRY OF BLOOD
1. The role of the liver in the metabolism of proteins and other nitrogen-containing
compounds. Diagnostic tests (urea, creatinine, plasma proteins), physiologic range,
clinical and diagnostic value. Plasma proteins: albumin, α1- and α2-globulins, βglobulins, γ-globulins. Acute phase proteins.
Role of liver:
- Synthesis of the major blood proteins (for transporting metabolites and metals,
immune function)
- Break down amino acids in foods so that they can be used to produce energy or make
carbohydrates or fats.
- Convert ammonia to a less toxic substance urea, which is released into the blood.
(detoxification)
Diagnosis tests (urea, creatinine, plasma proteins)
- Urea: measurement of urea in blood/urine.
+ Normal range of urea is 2.5 to 7.1 mmol/L.
+ The high level of urea can be caused by kidney, urinary system dysfunction or age,
high protein level intake, etc.
+ Low level of urea is due to liver dysfunction, but it is not common.
- Creatinine: measurement of creatinine in blood/urine.
+ Normal range: For adult men, 0.74 to 1.35 mg/dL (65.4 to 119.3 micromoles/L)
For adult women, 0.59 to 1.04 mg/dL (52.2 to 91.9 micromoles/L)
+ High level of creatinine shows kidney disease: diabetic nephropathy, or diabetic
kidney disease.
+ Low level of creatinine shows liver, muscles diseases due to age, pregnancy,
malnutrition.
- Plasma proteins: measurement of albumin, globulin, fibrinogen in the blood
+ Normal range: Albumin: 55% blood protein: 35–50 g/l
Globulin: 35% blood protein: 20–35 g/l
Fibrinogen: 6.5% blood protein
Plasma proteins: are synthesized in the liver except immunoglobulins (immune cells).
Almost plasma proteins are glycoprotein.
Function of plasma proteins:
- Transport substances: Albumin: fatty acids, bilirubin, calcium, drugs; b globulins:
transferrin: iron
- Osmotic regulation: albumin
- Catalytic function (enzymes): eg ceruloplasmin – ferroxidase
- Protective function (Ig combine w antigen to remove them, complement system
removes cellular antigen, …)
- Blood clotting (fibrinogen)
- Anticoagulant activity (thrombolysis): plasmin breaks down thrombin and dissolves the
clot
- Buffering capacity: maintain acid-base balance
Albumin: globular shape, relatively small size. Its polypeptide chain includes many
glutamate and aspartate residues
- Has 6 hydrophobic clefts: able to attach some low molecular weight
- Function: transport of hydrophobic molecules and maintaining oncotic pressure of the
blood. (metal: Ca, iron; free fatty acids; bilirubin; bile acid; hormone: steroids)
Globulins: globular shape, well soluble in water and water solutions
All synthesized in liver, and maybe small amount in other tissues.
Function
α1α1-antitrypsin
Serum trypsin inhibitor, protects tissues from enzymes of
globulins
inflammatory cells, especially neutrophil elastase
α1Inhibits proteases (cathepsin G) in neutrophils, chymases
antichymotrypsin in mast cells to protect tissues from damage by
proteolytic enzyme
α1-fetoprotein
Its appear means hepatocellular tumor, stomach,
intestinal tumors, …
orosomucoid
Carrier of basic and neutrally charged lipophilic
compounds
α2α2Inhibits proteases, coagulation; protein carrier (carry
globulins
macroglobulin
growth factors and cytokines
haptoglobin
Binds free hemoglobin and delivers it to the
reticuloendothelial cells
Complex Hb-Hp too large to pass through glomerulus ->
prevent loss of free Hb and Fe
Antioxidants (Fe2+->Fe3+)
ceruloplasmin
Copper carrier, plays role in iron and cooper metabolism,
antioxidant (Fe2+ -> Fe3+)
βtransferrin
Mediate the transport of iron through blood plasma,
globulins
antioxidants
hemopexin
Scavenging the heme released or lost by the turnover of
heme proteins, protects the body from the oxidative
damage that free heme can cause
Acute phase proteins: low specificity, appear in blood in case of injuries, fevers,
inflammation, stress. Their concentrations change during acute inflammatory response.
Functions:
- Antioxidant (ceruloplasmin), antiprotease (a2-macroglobulin), antibacterial (C-reactive
protein), hemocoagulant (fibrinogen)
Synthesized
Properties
Function
C-reactive
Liver
Consists of 5
Bind to C-polysaccharide of
protein
subunits
microbial wall, participate in
formation of primary immune
response
A1-antitrypsin
Liver
Serpin
Trypsin inhibitor, protects tissues
superfamily
from enzymes of inflammatory cells
Haptoglobin
Liver
Glycoprotein
Prevents removal of hemoglobin
from the body by kidneys, the
complex with hemoglobin is taken
up by reticuloendothelial system
and destroyed.
A2
Liver
Inhibits proteases, coagulation;
macroglobulin
protein carrier (carry growth
factors and cytokines
2. Role of the liver in carbohydrate metabolism: homeostasis of blood glucose, its
hormonal and metabolic regulation. Diagnostic tests (blood glucose, glucose tolerance
test), physiologic range, clinical and diagnostic value.
Role of liver in carbohydrate metabolism:
- Maintenance of blood glucose level. In the fed state, the liver takes up excess glucose
and stores it as glycogen or converts it to fatty acids. In the fasting state the
glycogenolysis and gluconeogenesis in the liver are major sources of glucose for the
body
- Hormonal regulation: insulin, glucagon
- Metabolic regulation:
+ Insulin: stimulates glucose intake of the cell to create energy or forming glucagon for
storage
Diagnostic test
- Blood glucose test: measure level of glucose level in the blood (not eat or drink 8
hours before the test)
+ Normal range: less than 140 mg/dL (7.8 mmol/L)
+ High glucose level: kidney disease, hyperthyroidism, pancreatitis, pancreatic cancer
+ Low glucose level: hypothyroidism, too much insulin, liver disease
- Glucose tolerance test: measure response of body to glucose (measure of glucose
level after glucose intake) to detect type 2 diabetes or prediabetes.
+ Normal range: less than 140 mg/dL (7.8 mmol/L)
+ A blood glucose level between 140 and 199 mg/dL (7.8 and 11 mmol/L) is considered
impaired glucose tolerance, or prediabetes.
+ A blood glucose level of 200 mg/dL (11.1 mmol/L) or higher may indicate diabetes.
3. Role of the liver in lipid metabolism: the main stages of the synthesis of
triacylglycerols, cholesterol, phospholipids, their hormonal and metabolic regulation;
lipoproteins formed in the liver, their structure and role; fatty liver disease, its causes;
diagnostic tests (cholesterol, TAG, HDL-Cholesterol, LDL-Cholesterol, atherogenic index),
physiologic range, clinical and diagnostic value.
Main stages synthesis of TAG, cholesterol, phospholipids
TAG
- Forming glycerol backbone (glycerol phosphate)
- Adding free fatty acid from fatty acyl CoA
Cholesterol: synthesized from acetyl CoA
- Synthesis of mevalonate from 3 acetate units
- Conversion of mevalonate to active isoprene units (isopentyl pyrophosphate and
dimethylallyl pyrophosphate)
- Polymerization of six isoprene units to form squalene
- Cyclization of squalene to yield four rings of steroid nucleus of lanosteroid.
Phospholipids
- Ethanolamine transform to phosphoethanolamine
- Phosphoethanolamine interacts with cytidine triphosphate forming CDP-ethanolamine
- CDP-ethanolamine condenses with diacylglycerol to yield phosphatidyl ethanolamine
- Forming phosphatidyl choline via methylation
- There are interrelated between phosphatidyl serine, choline and ethanolamine.
Lipoprotein forming in the liver
- Lipoprotein: a complex protein containing various lipids as a non-protein component
- Involved in synthesize of chylomicron, LDL, VLDL, HDL
- Structure: central part is lipid core (cholesteryl esters, triacylglycerol, non-polar lipids),
membrane is free cholesterol, phospholipid, and apoprotein
- Function: Carry fats (hydrophilic) to different tissues.
Fatty liver disease (hepatic steatosis): excess fat builds up in the liver
- Symptoms: no or few: tiredness or pain in abdomen
- Complications: cirrhosis, liver cancer, esophageal varices
- Treatment: lifestyle changes: diet, exercises
Diagnosis test
Cholesterol: measure of total amount of cholesterol (LDL, HDL, triglycerides) in the
blood
- Normal range:
+ LDL: 70 to 130 mg/dL (the lower the number, the better)
+ HDL: more than 40 to 60 mg/dL (the higher the number, the better)
+ Total cholesterol: less than 200 mg/dL (the lower the number, the better)
+ Triglycerides: 10 to 150 mg/dL (the lower the number, the better)
- High range of cholesterol number indicate the risk of heart disease, stroke and
atherosclerosis
TAG: measurement amount of TAG in VLDL in the blood helps to determine risk of
developing heart disease
- Normal range:
+ A normal fasting level is 150 milligrams per deciliter (mg/dL).
+ A borderline high level is 150 to 199 mg/dL.
+ A high level is 200 to 499 mg/dL.
+ A very high level is more than 500 mg/dL.
- Low triglyceride level maybe due to low fat diet, hyperthyroidism, malabsorption
syndrome, malnutrition
- High triglyceride level due to cirrhosis, diabetes, genetic factors, hyperlipidemia,
hyperthyroidism, kidney disease, pancreatitis
HDL-C: measure the level of good cholesterol in the blood
- Normal range:
+ Optimal levels of HDL cholesterol are over 60 milligrams per deciliter (mg/dL)
+ HDL levels that are below 40 mg/dL for men and 50 mg/dL for women indicate an
increased risk of heart disease.
- High level of HDL maybe due to C-reactive protein which is produced by liver when
inflammation occurs
- Low level of HDL due to lack of physical activities, smoking, …
LDL-C: measurement of LDC (bad cholesterol) in the blood to determine the risk of heart
disease
- Normal range: 70 to 130 mg/dL (the lower the number, the better)
- High level of LDL is caused by poor diet, obesity, lack of exercise, smoking, age and
diabetes. High cholesterol can cause chest pain, heart attack and stroke.
- Low level of LDL (less than 40 mg/dL) associated with cancer, hemorrhagic stroke,
depression, anxiety
Atherogenic index: is a novel index composed of triglycerides and high-density
lipoprotein cholesterol
- It is a marker of cardiovascular disease by predict the risk of atherosclerosis and
coronary heart disease
- Calculated by formula: log10 (TG/HDL-C)
- Range: involves to risk of CVD
+ 0.3 to 0.1 for low risk
+ 0.1 to 0.24 for medium
+ more than 0.24 for high risk
Apo-B/Apo-A1: predictor of cardiovascular risk
- Apo-A1 and ApoB are major apolipoprotein involved in lipid transport in the processes
causing atherosclerosis. ApoA1 is major protein in HDL while apoB indicates total
number of atherogenic particles.
- The higher the ratio the higher the cardiovascular risk. Normal range is from 0.19 to
2.60. At ratio 0.9 the risk of cardiovascular disease is 19.1%.
Lipoprotein A: measurement of level of lipoprotein A which is substance made of
protein and fat that carry cholesterol through bloodstream
- Normal range: less than 30 milligrams per deciliter (mg/dL).
- High lipoprotein mean the risk for heart disease
- Level of lipoprotein a is determined by genes and not affected by lifestyle or
medicines.
4. The role of the liver in digestion. The composition of bile and its role. The structure
and types of bile acids and the reactions of their synthesis. Causes of disorders of
synthesis and secretion of bile, consequences.
Role of liver in digestion:
- Produces bile
- Produces proteins and cholesterol
- Breakdown medications and alcohol
- Detoxification of ammonia
- Glycogen regulation and storage
- Synthesis of cholesterol
- Regulates and maintains level of blood glucose
Composition of bile:
Water (97.6%)
Substances (2.4%)
- Bile salts (50%) including bile salts
- Bile pigments (bilirubin) (2%)
- Cholesterol (4%)
- Phospholipids (40%)
- Inorganics substances: sodium, calcium, potassium, chloride, bicarbonate (4%)
Structure and types of bile acids. Reactions of synthesis
Bile acids:
- Primary: cholic acid, chenodeoxycholic acids
- Secondary: deoxycholic acid, lithocholic acid
Structure: precursor is cholesterol
Synthesis:
- Primary bile acids are synthesized in liver by transformation of cholesterol. They pass
into the bile and then stored and accumulated in gallbladder.
- Bile passes from gallbladder into duodenum. Secondary bile acids are products of
transformation of primary bile acids by enzyme of intestinal germs.
Disorder in synthesis and secretion of bile acids
Bile acids act as main pathway of breaking down and eliminating cholesterol from the
body. The failure in producing or secreting bile acids results in accumulation of
abnormal bile acids and other metabolites, and the excess amount of cholesterol in the
body can damage certain organ system
- Manifestations: malabsorption of fat and fat-soluble vitamin (cant absorb), yellow skin,
abdominal pain (upper right side of abdomen under rib cage), nausea, vomiting, fatigue,
loss of appetite, loss weight
- Diagnosis: blood test, liver function test
- Treatment: addition of missing bile acids
Microsomal oxidation
Responsible enzyme located in lipo-membrane of ER.
- After isolating small vessels obtained (microsome)
- Enzyme isolated from ER poses enzymatic activity called “microsomal mixed function
oxidase system”
- In the liver, NADPH are taken to the electron transport chain (flavoprotein 1) and then
it will activate the 2nd electron transport carrier cytochrome P450
- cytochrome P450 catalyses the hydroxylation (turn RH into ROH) and make toxic
substance hydrolyse to increase its solubility
- Toxic can cross the membrane, into the blood and eliminated by kidney filtration
- Using flavoprotein 1 (contain FAD and FMN)
NADPH+ is turned into NADPH by flavoprotein 1. At the same time, cytochrome p450
is activated and catalyzes the hydrolyse of RH into ROH using O2 and release H2O
5. The biotransformation of xenobiotics in the body. The role of the liver in the general
scheme of conversion of xenobiotics, its interaction with other organs. The scheme of
the process of microsomal oxidation. NADPH-dependent and NADH-dependent
pathways. Sources of NADH and NADPH, the components of the electron-transport
chains of microsomal system. The role of cytochrome P450. The substrates of
microsomal oxidation. Inducers and inhibitors of microsomal oxidation.
Biotransformation of xenobiotic compounds:
- Phase 1: addition or unmasking of a functional polar group. This results typically in a
relatively small increase in hydrophilicity and may cause metabolic activation. These
include enzymes such as cytochrome P450, alcohol dehydrogenase, hydrolase, epoxide
hydrolase, carbodyl esterase, etc
- Phase 2: conjugation with a small hydrophilic endogenous substance often, but not
always, to a functional group provided by a phase I reaction – thereby significantly
increasing hydrophilicity and facilitating excretion. Glutathione, sulfate, and glucuronic
acid serve for conjugation of xenobiotics.
Role of liver:
- Maintenance of blood glucose level
- The major site of fatty acid synthesis in human body
- The synthesis of ketone bodies during starvation
- The synthesis of plasma lipoproteins.
- The synthesis of bile acids from cholesterol
- The synthesis of the major of blood proteins
- Accumulation of some fat soluble vitamins (A,D) and iron (in complex with ferritin)
- Detoxification of endogenous toxic metabolites and xenobiotics
Interaction with other organs: Liver together with gallbladder, pancreas and intestines
work to digest, absorb and process food.
The liver's main job is to filter the blood coming from the digestive tract, before passing
it to the rest of the body. The liver also detoxifies chemicals and metabolizes drugs. As it
does so, the liver secretes bile that ends up back in the intestines. The liver also makes
proteins important for blood clotting and other functions.
NADPH-dependent and NADH dependent pathways
Sources of NADH and NADPH
- NADPH: pentose phosphate pathway (by glucose 6-phosphate dehydrogenase);
ferredoxin NADP+ reductase is major source of NADPH in photosynthetic plants.
- NADH: from transformation of pyruvate into acetyl CoA, glycolysis, citric acid cycle
Components of electron transport chain: cytochrome P450, NADOH (flavoproteins),
molecular O2, lipid membrane
Role of cytochrome P450
- Central role in phase 1 liver detoxification
- Reductive metabolism of xenobiotic substrate, endogenous compounds, steroid and
prostaglandins and fatty acids
- Assist in the synthesis vitamin D
- For hormone synthesis: effectively convert cholesterol into pregnenelone and then
converted into estrogen, testosterone, cortisol
- Necessary for assimilation of fat-soluble vitamins for synthesis of bile and bile acid
Substrate for microsomal oxidation
- Catechoalmine, histamine, ethanol, aldehyde, hypoxanthine, cyclohexene carboxylic
acid
Inducers: alcohol, cigarette, smoking, phenobarbitone hypnotic, phenytoin
Inhibitors: grape-fruit, cimetidine, antibiotics, antifungal.
6. The process of conjugation. The structure of UDP-glucuronic acid and
phosphoadenosine - phosphosulfate. Reactions of formation of conjugated bilirubin and
indican. Glycine conjugation, it’s role. Metabolism of ethanol. Alcohol dehydrogenase
and MEOS pathways. Toxicity of acetaldehyde. Causes of lactic acidosis, ketoacidosis
and hypoglycemia in alcohol intoxication.
The process of conjugation: makes the substrate more water soluble and capable of
being excreted by liver or biliary rout or renal route or blood and kidney filtration.
UDP-glucuronic: uridine diphosphate glucuronic acid: a UDP-sugar having alpha-Dglucuronic acid as the sugar component.
Phosphoadenosine-phosphosulfate: (PAPS) is a derivative of adenosine monophosphate
that is phosphorylated at the 3' position and has a sulfate group attached to the 5'
phosphate. It is the most common coenzyme in sulfotransferase reactions.
Reactions formation of conjugated bilirubin and indican.
Bilirubin:
(1) reticuloendothelial system (RES)
macrophages phagocytose old erythrocytes. Hemoglobin metabolism yields globin and
heme.
Pathway: heme → biliverdin (green-colored) → bilirubin (yellow-colored)
(2) bloodstream
albumin binds bilirubin and complex is carried to liver
bilirubin-albumin complex = indirect bilirubin (water insoluble)
(3) liver
hepatocytes take up bilirubin
hepatic microsomes conjugate bilirubin with glucuronic acid
conjugation via UDP glucuronyl transferase
enzyme is synthesized slowly after birth, sometimes causing newborn jaundice
conjugated bilirubin = direct bilirubin is water soluble
a portion of conjugated bilirubin is excreted in urine
remainder is secreted into bile and then into small intestine
Indican
Following absorption from the gut, indole together with sulfuric acid and potassium on
the decomposition of tryptophan is converted to 3-hydroxy indole (indoxyl or indican) in
the liver, where it is again then conjugated with sulfuric acid or glucuronic acid through
normal xenobiotic metabolism pathways. It is then transported to the kidneys for
excretion
Glycine conjugation:
- Catalyzed by glycine N-acyltransferase
Role:
- Important metabolic pathway responsible for maintaining adequate levels of free
coenzyme A (CoASH).
- Formation and toxicity of xenobiotic acyl CoAS
- Restoring levels of CoASH
Metabolism of ethanol
Oxidative pathway of alcohol metabolism: catalyzed by cytochrome P450, ADH (alcohol
dehydrogenase)
This oxidation process involves an intermediate carrier of electrons, nicotinamide
adenine dinucleotide (NAD+), which is reduced by two electrons to form NADH.
Nonoxidative pathway of alcohol is minimal. Alcohol is non-oxidatively metabolized in 2
pathways. 1st pathway leads to formation of fatty acid ethyl ester from reaction of
alcohol with fatty acids. (FAEEs). FAEEs are detectable in serum and other tissues after
alcohol ingestion and persist long after alcohol is eliminated. The role of FAEEs in
alcohol-induced tissue damage remains to be further evaluated.
The 2nd pathway results the formation of fat molecule containing phosphorus called
phosphatidyl ethanol. The product of this reaction, phosphatidyl ethanol, is poorly
metabolized and may accumulate to detectable levels following chronic consumption of
large amounts of alcohol, but its effects on the cell remain to be established. However,
the formation of phosphatidyl ethanol occurs at the expense of the normal function of
PLD, namely to produce PA, resulting in inhibited PA formation and disruption of cell
signaling.
ADH: involved to the major pathway of oxidative metabolism of ethanol in the liver.
ADH constitutes a complex enzyme family, and, in humans, five classes have been
categorized based on their kinetic and structural properties.
Metabolism of ethanol with ADH produces acetaldehyde, a highly reactive and toxic
byproduct that may contribute to tissue damage. Alcohol oxidation generates a highly
reduced cytosolic environment in liver cells (i.e., hepatocytes). In other words, these
reactions leave the liver cells in a state that is particularly vulnerable to damage from
the byproducts of ethanol metabolism, such as free radicals and acetaldehyde.
Toxicity of acetaldehyde:
Acetaldehyde enters your blood, damaging your membranes and possibly causing scar
tissue. It also leads to a hangover, and can result in a faster heartbeat, a headache, or
an upset stomach. The brain is most affected by acetaldehyde poisoning. It causes
problems with brain activity and can impair memory.
Causes of lactic acidosis, ketoacidosis and hypoglycemia in alcohol toxification
Lactic acidosis: heart disease, severe infection, HIV, chronic alcoholism, intense
exercise, or physical activity
Ketoacidosis: deficiency of insulin, illness, infection
Hypoglycemia in alcohol in toxification: Alcohol consumption causes an increase in
insulin secretion, which leads to low blood sugar
7. The structure and synthesis of heme. Formation of porphobilinogen, scheme of
synthesis of proto-porphyrin IX and its transformation into a heme. Role of
ferrochelatase. Regulation of the process. Disorders of the synthesis of heme and
hemoglobin: porphyria and thalassemia.
Structure of heme:
- Porphyrin nucleus
- 4 pyrrole rings (tetrapyrrole)
- Bridges – methine (CH)
- Side chain: 8: methyl 4, vinyl CHCH2 2, propionic acid CH2CH2COOH 2
Heme synthesis: major site is in liver and erythrocyte-producing cells of bone marrow
Transformation into heme
Role of ferrochelatase:
- Catalyzes the insertion of ferrous iron into protoporphyrin IX, yielding heme.
- Decreased values of ferrochelatase activity in all tissues are a characteristic of patients
with protoporphyria.
Regulation of the process heme synthesis:
- ALA synthase is regulated by repression mechanism
- Heme inhibits the synthesis of ALA synthase by acting as co-repressor (negative
inhibition)
- ALA synthase is allosterically inhibited by hematin. Excess Fe2+ is oxidized into Fe3+
(ferric), forming hematin
Heme protein: cytochrome, myoglobin
Porphyria: group of genetic disorders of producing porphyrin which is essential for heme
function in human body
Types: hepatic, erythropoietic or both
- Hepatic forms of the disorder caused by liver problem and associated with abdominal
pain and central nervous system problems
- Erythropoietic forms caused by red blood cells problem and associated with light
sensitivity
Symptoms: hypersensitivity to the light, acute attach, vomiting, muscle weakness,
abdominal pain, etc
Treatment: barbiturates (reduce ALA synthase)
Diagnosis: UV fluorescence is the best technique
Thalassemia: inherited disease which cause hypoproduction of hemoglobin chain
- Alpha thalassemia: reduced in production of alpha chains of hemoglobin
+ One mutated gene, you'll have no signs or symptoms of thalassemia. But you are a
carrier of the disease and can pass it on to your children.
+ Two mutated genes, your thalassemia signs and symptoms will be mild. This
condition might be called alpha-thalassemia trait.
+ Three mutated genes, your signs and symptoms will be moderate to severe.
- Beta thalassemia: reduced in production of beta chains of hemoglobin (common)
Risk factors: family history of thalassemia, certain ancestry (from African Americans or
Southeast Asian)
Complications: iron overload, enlarged spleen, slow growth rate, heart problems, bone
deformities, dark urine, yellow or pale skin
Diagnosis: blood test: test for anemia and abnormal hemoglobin
Treatment: blood transfusion, bone marrow transplantation, medications, spleen or
gallbladder removal surgery.
8. The degradation of hemoglobin and formation of bilirubin in the reticuloendothelial
system. Transport of bilirubin to the liver. Stages of bilirubin metabolism in the liver. The
role of the enzyme UDP-glucuronyl transferase. Steps of bilirubin metabolism in the
intestine.
Degradation of hemoglobin
- The hemoglobin is degraded (by reticuloendothelial cell phagocytosis) into globin - the
protein part and heme. The heme initially breaks apart into biliverdin, a green pigment
which is rapidly reduced to bilirubin, (heme oxygenase) an orange-yellow pigment.
Bilirubin formation in reticuloendothelial system
- In the first oxidation step, biliverdin is formed from heme through the action of heme
oxygenase, the rate-limiting step in the process, releasing iron and carbon monoxide.
- Next, water-soluble biliverdin is reduced to bilirubin. Because of its hydrophobic
nature, unconjugated bilirubin is transported in the plasma tightly bound to albumin.
Binding to other proteins and erythrocytes also occurs.
Approximately 75% of bilirubin is derived from hemoglobin, but degradation of
myoglobin, cytochromes, and catalase also contributes.
Bilirubin transportation to the liver
Because unconjugated bilirubin has hydrophobic nature, it tightly bound to albumin for
transportation from plasma to the liver
In the liver, bilirubin is removed from albumin, taken up at sinusoidal surface of
hepatocyte by facilitated transport system and then concentrated.
Stages of bilirubin metabolism in the liver
- Unconjugated bilirubin (hydrophobic) is transported to liver by binding to albumin
- In the liver, glucuronic acid is added to unconjugated bilirubin (catalyzed by glucuronyl
transferase), forming conjugated bilirubin which is water soluble and can be excreted
into duodenum in bile
Role of UDP-glucuronyl transferase: catalyzes reaction forming conjugated bilirubin by
glucuronic acid as well as phase 2 of biotransformation in detoxification.
Steps of bilirubin metabolism in the intestine
- Conjugated bilirubin is excreted through bile to the intestine
- Bacterial action (deconjugation and reduction) forms bilinogens (stercobilinogen,
mesobilinogen and urobilinogen). About 20% of the urobilinogen is reabsorbed daily
from the intestine to enter enterohepatic circulation to get re excreted into the
intestinal lumen (enterohepatic circulation). A small fraction of urobilinogen enters the
systemic circulation and gets filtered at the glomerulus and excreted in urine.
- Urobilin is excreted by the kidneys to give urine its yellow color and stercobilin is
excreted in the feces giving stool its characteristic brown color.
9. Jaundice, types, causes, laboratory criteria. Neonatal jaundice. Pathological jaundice.
Types
Hemolytic jaundice
Obstructive
Parenchymal
jaundice
jaundice
Causes
Hyperdestruction of
The blockage of
jaundice caused by
hemoglobin leads to
bile flow from the the liver cell
excess level of
liver to the
damage and can be
bilirubin in the blood: intestine causes
cured by
the accumulation
medication
- Hemolysis
of the bile in
- Resorption of
gallbladder,
- Hepatitis A, B, C, E
hematoma
forming gallstone. - Drug toxicity
(poisoned liver)
- Minor/ major
incompatibility
- Massive transfusion
Laboratory criteria
Elevate lactate
dehydrogenase in the
blood
Surgical treatment
is required.
- Parasite,
common bile duct
stone
Bilirubin level in
the blood
- Cirrhosis
- Paracetamol
overdose
Biopsy for checking
damage of the liver
Neonetal jaundice: jaundice appears in newborn
Physiological jaundice: appears after 24h of birth
- Transient jaundice: deficiency of UDP glucuronyl transferase
- Breast milk jaundice: hyperbilirubinemia caused by mother’s milk estrogen inhibits
UDP glucuronyl transferase
Pathological jaundice: appears within 24h of birth
Appears within 24 hours of birth
Hemolytic jaundice: result of hemolysis or an accelerated break down of RBCs lead to
increasing production of bilirubin
Causes: excess production of bilirubin due to excess break down of hemoglobin
Biochemical basis for the use of phenobarbital
The major effect of phenobarbital is to increase hepatic UDP glucuronyl transferase
(UGT) activity and the conjugation of bilirubin, apart from possibly enhancing hepatic
uptake of bilirubin.
Hereditary disorders excretion of bilirubin
Gilbert syndrome: decreased in UDP glucuronosyltransferase activity
- Unconjugated hyperbilirubinemia accumulated in the liver causing the yellowish of the
skin, white of the eyes
- Jaundice may be triggered by stress such as exercises, menstruation, not eating.
- Diagnosis based on level of unconjugated bilirubin in the blood without signs of liver
problems or RBCs breakdown
- Treatment: for significant jaundice: phenobarbital
Dubin-Johnson syndrome: defecting bilirubin transport out of the liver because of
mutated canalicular membrane carrier
- Manifestations: conjugated hyperbilirubilin, black liver on gross pathology (Deposition
of conjugated bilirubin which is giving black to the liver)
- Asymptomatic, no treatment needed
- Diagnosis: unusual ratio between byproducts of heme biosynthesis (coproporphyrin III
and I), autopsy (liver have dark pink or black colour)
Crigler-Najjar syndrome: absence of UDP glucuronyl transferase acitivity
- Types:
+ Type I: fatal within 2 years of birth, presence of jaundice, kernicterus, unconjugated
hyperbilirubinemia
+ Type II: nonfatal, presence of jaundice and unconjugated hyperbilirubinemia
- Symptoms: yellowish of skin, white of the eyes
- Diagnosis: patient history, blood test to reveal high level of unconjugated bilirubin in
the absence of RBC degeneration increasing, bile analysis, urine analysis, molecular
genetic testing.
- Treatment: plasmapheresis and phototherapy (type I) and thenobarbital (type II)
Obstructive jaundice: blockage the flow bile out of the liver
Cystic fibrosis: mutation on chromosome 7, results in defective membrane Cl- channel
- Mutation in Cl- channel causes defective chloride and water transportation in epithelial
cells. This result secretion and trapping of thick mucus plugs in lungs, liver and pancreas
- Manifestations: chronic lung disease, pulmonary infection, steatorrhea,
malabsorption, abdominal pain
- Diagnosis: High Cl- concentrations in sweat test, increasing in ratio of residual volume
to total lung capacity.
- Treatment: lung transplantation, bronchodilators, technique for clear airway
secretions.
Niemann-Pick disease: deficiency of sphingomyelinase (for converting sphingomyelin to
ceramide during glycosphingolipid catabolism)
- Harmful quantities of lipids accumulate in the brain, spleen, liver, lungs, and bone
marrow. Neurological symptoms may include ataxia (lack of muscle control during
voluntary movements such as walking), loss of muscle tone, brain degeneration,
increased sensitivity to touch, spasticity (stiff muscles and awkward movement), and
slurred speech, difficulties in swallowing, eye paralysis, learning problem, enlargement
of spleen and liver
- Types
+Type A: present in first 6 months of life with progressive CNS deterioration, seizures,
failure to thrive
+ Type B: present in teenager with milder nervous system involment
+ Type C: present in childhood with ataxia, dystonia. Patient can develop mental
retarded, hepatosplenomegaly, progressive intestinal pulmonary disease
- Diagnosis: reticular infiltrative pattern on chest x-ray film
- Treatment: hepatic or bone marrow transplantation, enzyme therapy
Hypoplasia of the biliary tract: lack of development of biliary tract
- In liver biopsy finding reduced amount of bile ducts
- Causes: age, postoperative bilirubin level, hispathology of liver, cholangitis
- Manifestation: jaundice: yellowish of the skin, white of the eyes
- Diagnosis: cholangiography
- Treatment: decompression and irrigation of biliary tract
10. Metabolism of iron: daily requirement, nutritional sources, mechanism of
absorption, transport in blood, mechanism of transport across the cell membrane,
storage form. Iron- containing proteins. Regulation of iron metabolism. The role of
hepcidin and cytokines.
DAILY REQUIREMENT
The total body iron content is 3.5 to 4 g; 75% of which is in blood, the rest is in the liver,
bone marrow and muscles.
In blood: About 75% of total iron is in hemoglobin, and 5% is in myoglobin and 15% in
ferritin.
Requirement of Iron (ICMR)
Iron loss is 1 mg/day
Infants up to 4 months
0.5 mg
Infants 5-12 months and children
1 mg
Menstruating women
3 mg
Pregnancy
3-4 mg
Adult men and postmenopausal women
1mg
Sources of Iron
•
liver (but avoid this during pregnancy)
•
red meat
•
beans, such as red kidney beans, edamame beans and chickpeas
•
nuts
•
dried fruit – such as dried apricots
•
fortified breakfast cereals
•
soy bean flour
Summary of Iron Absorption
- Free iron from ingestion in the intestines is reduced from the ferric (Fe3+) to the
ferrous (Fe2+) state on the apical surface of intestinal enterocytes and transported into
the cells through the action of the divalent metal transporter (DMT1).
- Intestinal uptake of heme iron occurs through the help of heme transporter (HCP1).
The iron is then released within the enterocytes by the enzyme heme oxygenase.
- The iron can be temporarily stored within enterocytes bound to ferritin. Iron is
transported across the basolateral membrane of intestinal enterocytes into the blood,
through the action of the transport protein ferroportin.
- Iron which are temporarily stored in enterocytes loss can take place due to shedding
of epithelial cells
Iron Transport in Blood and Uptake by Cells
Iron in the mucosal cells through DMT1, HCP1 and ferroportin
The iron (Fe3+) first is reduced to ferrous state (Fe2+) by ascorbic acid entering the
mucosal cells by absorption. Fe2+ then combines with apoferritin to form ferritin which
is the temporary storage form of iron. From the mucosal cells, iron may enter the blood
stream (which mainly depends on the body needs) or lost when the cells are
desquamated.
Transport of Fe in the plasma.
The iron liberated from the ferritin of mucosal cells enters the plasma in ferrous state.
Here, it is oxidized to ferric form (Fe3+) by a copper-containing protein-ceruloplasmin
(ferroxidase) which possesses ferroxidase activity. Another cuproprotein ferroxidase II
also helps for the conversion of Fe2+ to Fe3+. Ferric iron then binds with a specific iron
binding protein, namely transferrin or siderophilin (a glycoprotein). Each transferrin
molecule can bind with two atoms of ferric iron (Fe3+).
Storage of Iron
- The storage form is ferritin. It is seen in intestinal mucosal cells, liver, spleen, and bone
marrow.
+ The apoferritin has a molecular weight of about 440 kilo Daltons. It has 24 subunits. It
can take up to 4,000 iron atoms per molecule. Ferritin contains about 23% iron.
+ Normal plasma contains very little ferritin. Ferritin in plasma is elevated in iron
overload. Ferritin level in blood is an index of body iron stores.
+ Ferritin is an acute phase reactant protein that is elevated in inflammatory diseases.
Estimation of ferritin is also indicated in chronic kidney disease to assess the extent of
anemia.
- Aging cells will release iron with the help of a copper containing protein called
hephaestin, which has ferroxidase activity.
- Hemosiderin is also a storage form of iron, but it is formed by partial deproteinization
of ferritin by lysosomes and are found as aggregates in tissues like liver, spleen and
bone marrow. It is more insoluble than ferritin, and iron is more slowly released.
Iron containing proteins: haemoglobin, myoglobin, cytochrome, catalase, glutathione
peroxidase, thyreoperoxidase, myeloperoxidase
Regulation of Absorption by Four Mechanisms
i. Mucosal regulation: Regulation by mucosal block theory
- Absorption of iron needs divalent metal ion transporter (DMT) and ferroportin.
- Synthesis of both these proteins is down regulated by hepcidin, a peptide secreted by
the liver when body iron reserves are adequate. If there is hypoxia or anemia, the
synthesis of hepcidin is reduced; so ferroportin synthesis will increase.
- Hepcidin It is produced by liver cells and is involved in killing bacteria. It is coded in
HAMP gene on chromosome 19. Its main action is on iron metabolism. Hepcidin
decreases surface expression of the ferroportin, which is responsible for moving iron
across cell membranes. Hepcidin production is increased by high iron stores and by
inflammation.
ii. Storage regulation: As body iron store falls, the mucosa is signaled to increase
absorption.
iii. Erythropoietic regulation: In response to anemia, the erythroid cells will signal the
mucosa to increase iron absorption. This signal may be erythropoietin from kidney.
iv. Reciprocal relationship between synthesis of ferritin and transferrin receptor (TfR).
- In case of high concentration of iron, the iron bound to the IRE-BP (Iron-Responsive
Element Binding Protein) prevents it to bind Iron-Responsive Element (IRE) of both
mRNA molecules. So, mRNA for ferritin is translated and ferritin is synthesized. But
mRNA for TfR is degraded, resulting in reduced TfR protein synthesis. Thus, when iron
levels are high, ferritin is synthesized to store iron. At the same time, there is no
requirement for further uptake of iron, so the TfR is not synthesized. This is a good
example of control of protein synthesis at the translational level. The reverse occurs
when iron status is low- ferritin mRNA is not translated, but TfR m RNA is translated.
cytokines derived from T cells and monocytes regulate cellular iron homeostasis by
affecting the expression of proteins involved in the uptake and storage of the metal.
Proinflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin-1 (IL-1),
or interleukin-6 (IL-6) directly stimulate the transcription and translation of the major
iron storage protein ferritin, the latter by interfering with a so-called acute phase box.
cytokines influence the posttranscriptional control of iron homeostasis by modulating
the binding affinity of iron regulatory proteins (IRP-1 and IRP-2) to specific RNA stem
loop structures, termed iron responsive elements (IREs), which are found within the 5ʹ
untranslated region of ferritin mRNA and within the 3ʹ untranslated region of transferrin
receptor (TfR) mRNA, the surface protein involved in the uptake of transferrin-bound
iron. Activation of IRP binding to IREs is controlled by intracellular iron availability, with
IRP binding affinity being high when intracellular iron concentrations are low, thus
resulting in stabilization of TfR mRNA while ferritin translation is reduced.
11. Deficit of iron and excess of iron in organism: causes, biochemical effects and clinical
manifestations. Hemochromatosis. Iron deficiency state.
Deficit of iron
Causes of iron deficit and biochemical effects
1. Nutritional deficiency of iron
2. Lack of absorption: Subtotal gastrectomy and hypochlorhydria
3. Hookworm infection: One hookworm will cause the loss of about 0.3 mL of blood
per day. Calculation shows that about 300 worms can produce a loss of 1% of total
body iron per day
4. Repeated pregnancies: About 1g of iron is lost from the mother during one
delivery
5. Chronic blood loss: Hemorrhoids (piles), peptic ulcer, menorrhagia
6. Nephrosis: Haptoglobin, hemopexin and transferrin are lost in urine, along with
loss of iron
7. Lead poisoning: Iron absorption and hemoglobin synthesis are reduced. In turn,
iron deficiency causes more lead absorption. It is a vicious cycle.
Iron deficiency is characterized by microcytic hypochromic anemia. Anemia results when
hemoglobin level is less than 12 g/dL.
Clinical manifestations
- When the level is lower than 10 g, body cells lack oxygen and patient becomes
uninterested in surroundings (apathy). Since iron is an important constituent of
cytochromes, their deficiency leads to derangement in cellular respiration and all
metabolic processes become sluggish.
- Prolonged iron deficiency causes atrophy of gastric epithelium leading to achlorhydria,
which in turn causes lesser absorption of iron, aggravating the anemia. Similar atrophy of
epithelium in oral cavity and esophagus causes dysphagia termed Plummer- Wilson
syndrome, which is a known precancerous condition.
- Very chronic iron deficiency anemia will lead to impaired attention, irritability, lowered
memory and poor scholastic performance. Anemia and apathy go hand in hand.
Excess of iron in organism: Iron excess is called hemosiderosis (accumulation in tissue
normally do not contain iron) or hemochromatosis (deposition of iron in liver, spleen,
pancreas, and skin)
Causes
When RBC die, it releases iron which becomes hemosiderin. Hemosiderin together with
ferritin are storage form of iron in body tissue. Accumulation of hemosiderin can cause
hemosiderosis. Causes:
- Bleeding within an organ or area of tissue
- RBCs break down within bloodstream
- Repeated blood transfusion
Biochemical effects
Free iron is a pro-oxidant – the opposite of an antioxidant – and may cause damage to
cells.
Clinical manifestation:
- In the lung, iron overload decreases blood oxygen saturation due to pulmonary
hypertension, autoimmune condition, chronic lung infections
- In the kidney: hemolytic anemia, autoimmune conditions
- Difficulty breathing, fatigue, shortness of breath, slow growth in children
Hemochromatosis Iron overload in liver, spleen, pancreas, joints, and skin
- In the liver, hemosiderin deposit leads to death of cells and cirrhosis.
- Pancreatic cell death leads to diabetes.
-Deposits under the skin cause bronze or grey discoloration
Types:
- Primary: genetic disorder
- Secondary: associated with certain anemia, liver disease and getting a lot of blood
transfusion
Symptoms: pain in joints, tiredness, skin has bronze or grey colour, foggy memory, etc
Diagnosis: based on symptoms, check family history, stethoscope, blood test (transferrin
saturation, serum ferritin), liver biopsy, MRI
Treatment: removing blood from the body (to drive iron level to normal)
12. Role of blood in transport of oxygen. Scheme of reactions occurring in the
erythrocyte in capillaries of lungs and in peripheral tissues. Transport of carbon dioxide.
The role of carbonic anhydrase. An influence of processes in erythrocytes on
concentration of bicarbonate in plasma. The mechanism of binding of the heme of
hemoglobin with oxygen, a role in the regulation of acid-base balance.
Role
Blood collects oxygen in respiratory organs and releases it in tissues in order to generate
the energy necessary for cell survival by producing energy.
Reactions occur in erythrocyte in capillaries of lungs and peripheral tissues: due to
passive difussion of partial pressure differences
- In the lungs: Oxygen diffuses from the alveoli into the pulmonary capillary blood
because PO2 in alveoli (104mmHg) greater than PO2 in pulmonary capillary blood
(40mmHg)
- In the body tissues, higher PO2 (100mmHg) in the capillary blood causes the diffusion
of oxygen to surrounding cells (PO2=40mmHg)
Scheme of reactions occurring in the erythrocyte:
Transport of carbon dioxide: 3 ways
- Dissolved CO2 (10%)
- Bound to hemoglobin (20%)
- Bicarbonate HCO3- (70%)
1. Dissolved CO2
- CO2 is more soluble in blood than oxygen.
2. Bound to hemoglobin
- The binding of CO2 to hemoglobin is reversible, forming carbaminohemoglobin. When
it reaches the lungs, the CO2 can freely dissociate from hemoglobin and be expelled
from the body
3. Bicarbonate
- Carbonic anhydrase converts CO2 into carbonic acid (H2CO3). Carbonic acid is
unstable, forming bicarbonate ion and hydrogen ion immediately. Then CO2 can
continue to dissolve into the blood, down its concentration gradient
- Hemoglobin can bind to free H+ ion, limiting shift in pH. The newly synthesized
bicarbonate ion is transported out of the RBC into liquid component in exchange for Clion (Chloride shift).
- When blood reaches the lung, bicarbonate ions are transported back into RBC in
exchange for Cl-. The H+ ion dissociates from hemoglobin and binds to bicarbonate ion.
Carbonic acid is produced then converted back to CO2 through action of enzyme of CA.
CO2 is expelled in the lungs during exhalation
Role of carbonic anhydrase
- Providing further dissociation of CO2 into the blood.
- Converting back HCO3- and H+ ions into CO2 and H2O for regulate acid-base balance
and exhalation of CO2.
Influence of bicarbonate concentration in plasma to erythrocyte
- Bicarbonate ion serves as regulator of red blood cell sodium level. The plasma
electrolyte changes causes changes in arterial pressure elevation (by trigger action
potential)
The mechanism of binding of the heme of hemoglobin with oxygen:
Hemoglobin is a protein found in RBC made of four subunits: two alpha subunits and
two beta subunits. Each subunit has a central heme group that contains iron and binds
one oxygen molecule, allowing each hemoglobin molecule to bind four oxygen
molecules. Molecules with more oxygen bound to the heme groups are brighter red.
Binding of first molecule of oxygen to hemoglobin will increase affinity to the 2nd and
then the 3rd and 4th.
A role in the regulation of acid-base balance:
As blood near the lungs, the carbon dioxide concentration decreases, causing an
increase in pH. The increases hemoglobin’s affinity for oxygen through the Bohr effect,
causing hemoglobin to pick up oxygen entering the blood from the lungs so it can
transport to the tissues. In the tissues, by contrast, the increasing CO2 concentration
drives a decrease in pH, which helps force hemoglobin to dump the oxygen it’s carried
from the lungs, so the cells can use it to break down sugars for energy. The pH-mediated
change in affinity for oxygen helps hemoglobin act like a shuttle that pick-up oxygen in
the lungs and deposits it in the tissues where it will be needed.
13. Acid-base status of the blood. The role of constant concentration of H+ ions. Sources
of H+ ions in cells. Key indicators of acid-base balance (pH, pCO2, pO2, HbO2, SO2, anion
gap), normal range. Effect of liver, secretion of stomach, pancreas and intestine on acidbase status of the body.
Acid base status of the blood:
Normal pH: 7.35 - 7.45 (slightly alkaline)
Maintenance of blood pH is important in hemostatic mechanism of body for normal
function of proteins, hormones, and receptors.
Role of constant concentration of H+ ions:
• Maintaining activity of enzymatic and transport proteins
• Non-specific protection of skin epithelium
• Negative charge of outer surface of erythrocyte membrane, with a decrease in
charge of the membranes, RBCs begin to aggregate, forming “coin columns”
which increases the viscosity of the blood
• Solubility of inorganic and organic molecules (Ca2+, Mg2+, oxalic acid and uric
acid)
• Formation of electrochemical gradient in mitochondrial membrane at the proper
level and activity of catabolic processes
Hydrogen index: its value determines the diagnosis of acidosis or alkalosis
Sources of hydrogen ions in the body:
1. In the reactions of aerobic metabolism of glucose, amino acids and fatty acids,
CO2 molecules are constantly formed with participation of carbonic anhydrase
enzyme. It reacts with water and forms carbonic acid which weakly dissociates
into H+ ion and bicarbonate ion. This method of proton production is
characteristic of almost all cells and occurs during aerobic metabolism, when
oxidative decarboxylation of pyruvate and TCA are active.
2. Aerobic glucose metabolism in which lactic acid appears:
Glucose
lactic acid
lactate + H+
- Occurs especially during intense muscular workout
- Accumulation of lactic acid is encountered with insufficient O2 supply to cells- leading
to anemia, shock, thrombosis and respiratory failure, etc.
- Deficiency of iron and copper in the composition of the respiratory chain enzymes also
play a role.
3. Metabolism of sulfur containing A.a and other compounds leads to appearance
of sulfuric acid which dissociates into sulfate ion and H+ ion
Methionine
H2SO4
SO4 (2-) + 2H+
Cysteine
H2SO4
SO4(2-)+ 2H+
4. Under certain conditions (starvation, Type 1 Diabetes mellitus) ,ketone
bodies(Acetoacetic acid, Hydroxybutyric acid ) enter the bloodstream
Fatty acids
acetyl SCoA
Acetoacetic acid
Acetoacetate+ H+
Fatty acids
acetyl SCoA
hydroxybutyric acid
Hydroxybutyrate + H+
5. Incase of poisoning with organic compounds, oxalic acid and formic acid can
serve as a source of H+ ions during metabolism of ethylene glycol and methanol.
Key indicators of acid-base balanace (pH, pCO2, pO2, HbO2, SO2, anion gap)
pH measuring the H+ ion level. It’s kept at 7.35-7.45 for homeostasis.
PCO2-The partial pressure of carbon dioxide in the blood of the capillary is about 45 mm
Hg, whereas its partial pressure in the alveoli is about 40 mm Hg.
PO2-The partial pressure of oxygen in the alveoli is about 104 mm Hg, whereas its
partial pressure in the blood of the capillary is about 40 mm Hg. This large difference in
partial pressure creates a very strong pressure gradient that causes oxygen to rapidly
cross the respiratory membrane from the alveoli into the blood.
HbO2- 100%
Hb molecule is composed of 4 subunits (α1,α2,β1,β2). The binding of 1st,2nd, 3rd oxygen O2
will increase affinity of Hb to 2nd,3rd,4th O2 molecules respectively.
SO2- Short-term exposures to SO2 can harm the human respiratory system and make
breathing difficult. People with asthma, particularly children, are sensitive to these
effects of SO2.
•
•
•
•
•
•
SO2 safe levels
Good (0–0.1 ppm) No cautionary statement.
Moderate (0.1–0.2 ppm) ...
Unhealthy for Sensitive Groups (0.2–1.0 ppm) ...
Unhealthy (1.0–3.0 ppm) ...
Very Unhealthy (3.0–5.0 ppm) ...
Hazardous ( > 5.0 ppm)
Anion gap - The anion gap is the difference between primary measured cations (sodium
Na+ and potassium K+) and the primary measured anions (chloride Cl- and bicarbonate
HCO3-) in serum. A high anion gap value means that your blood is more acidic
than normal.
Normal results generally fall between 3 and 10 milliequivalents per liter.
Effect of liver, secretion of stomach, pancreas, and intestine on acid-base status of the
body.
Liver influences
Carbohydrate metabolism
- Glucose can be stored as glycogen within the liver or it can be converted into more
stable storage form as triglycerides.
- End products are volatile acids: carbonic acid. Then CO2 is formed under catalyzed of
carbonic anhydrase and eliminated through the lungs
- Both acidosis and alkadosis decrease insulin secretion. Therefore, they cause fasting
hyperglycemia and decrease glucose tolerance
Amino acid metabolism
- Proteins break down in the intestine and absorbed as amino acids which then reach
liver. There they may be utilized to form proteins of different kinds. Some of them are
produced only in the liver, examples are albumin, alpha and beta globulins and
coagulation factors
- Acid base balance and amino acid metabolism are intimately related.
+ Glutamine is involved in renal ammonia genesis, a process related to acid excretion
+ Metabolism of serine, glycine, and branched chain aa influence by acid base balance
+ Basic cations (from lysine, arginine and histidine) yield neutral end products plus a
proton; sulfur (from methionine and cysteine) is acidogenic because they generate
sulfuric acid when oxidized; anionic aa (aspartate and glutamate) consume acid when
oxidized and reduce the acid load of the diet
- Products: nonvolatile acids: sulfuric acid, phosphoric acid and others. They are strong
acids that give up their H+ ion and secreted into urine by kidney.
Lipid Metabolism
- Fatty acids will be catabolized to release acetyl CoA. It may be used in the TCA cycle
and Electron transport chain to release energy or to act as a source of carbon for fatty
acid and cholesterol synthesis in healthy individuals. A small portion of acetyl CoA is
converted to ketone bodies (acetone, acetoacetic acid and beta hydroxybutyric acid).
The protein parts of the lipoproteins, apoproteins are synthesized by the liver only.
- End products are volatile acids: carbonic acid. Then CO2 is formed under catalyzed of
carbonic anhydrase and eliminated through the lungs
Bilirubin Metabolism
The heme present in the hemoglobin and other proteins/enzymes (e.g. cytochromes)
are eliminated only through liver
Secretion while on acid-base status
In stomach:
The gastric mucosa has different types of cells: (a) The mucous secreting mucus on the
surface of epithelial cells (b) The parietal cells which secrete acid (c) The chief cells
secrete enzymes.
The HCl secreted by the parietal cells creates a pH 0.8. Regulated by gastric and
histamine
In pancreas:
The fluid is alkaline and contains bicarbonate and enzymes. This secretion is under the
control of the hormones, secretin, and cholecystokinin. Secretin is produced under the
stimulation of gastric HCl. Secretin produces a secretion with high bicarbonate content.
Gastrin stimulates production of cholecystokinin (CCK), which in turn produces
pancreatic secretion rich in enzymes. The major enzymes present in pancreatic juice are
amylase, lipase, and proteolytic enzymes (trypsin, chymotrypsin, carboxypeptidase,
elastase) as their zymogens.
In intestines:
The start of the intestinal phase of secretion is mediated by the entry of chyme into the
small intestines.
Where the remaining 10% of acid that wasn’t secreted in the stomach and pancreas is
secreted.
Intestine distention and amino acids stimulate this kind of secretion in the intestines.
14. The chemical mechanisms of regulation of acid-base status. The buffer system of
blood (phosphate buffer, proteins, bicarbonate buffer, hemoglobin). Physiological
compensation of acid-base imbalance - the role of the lungs, kidneys and bones, the
mechanisms.
The chemical mechanism of regulation of acid base status: regulate level of H+ ion in the
plasma
Buffer system of blood
Bicarbonate Buffer System
- This buffer system operates both in the lungs and kidneys.
- This is the major extracellular buffer system.
- Lungs can decrease the carbonic acid by blowing out the CO2 and leaving water
behind.
- Kidneys can reabsorb HCO3- or regenerate new HCO3- from CO2 and water.
- Both the systems are very efficient because:
+ HCO3– is easily reabsorbed or regenerated by the kidneys.
+ The lungs adjust acid concentration.
Compensation of the pH is done as:
- The respiratory system compensates pH by decreasing or increasing CO2 by changing
the rate of respiration.
- The renal system produces more acidic or more alkaline urine.
Phosphate buffer system:
- Phosphates are found in 2 forms: H2PO4- or HPO4 2- together with Na+ ions.
- It’s the most important for intracellular buffer system and in developing urine
Protein buffer system: The plasma proteins and hemoglobin together constitute the
protein buffer system of the blood. The buffer ability depends on the bipolar of amino
acid residues: NH3+ and COOHemoglobin as a buffer
- Hemoglobin (Hb) is the best intracellular buffer system, and it combines with H+ and
forming HHb and CO2, forming the HHbCO2 complex.
- Hb combining with H+ ion becomes weak acid.
- Venous blood Hb is a better to buffer system than arterial blood Hb.
Physiological compensation of acid-base imbalance - the role of the lungs, kidneys and
bones.
The lungs: Releases carbon dioxide from the lungs. As carbon dioxide accumulates in the
blood, the pH of the blood decreases (acidity increases). As amount of carbon dioxide
exhaled, and pH of the blood increases as breathing becomes faster and deeper. By
adjusting the speed and depth of breathing, the brain and lungs can regulate the blood
pH minute by minute.
The kidneys: their cells reabsorb bicarbonate HCO3− from the urine back to the blood
and they secrete hydrogen H+ ions into the urine. By adjusting the amounts reabsorbed
and secreted, they balance the bloodstream pH. This adjustment takes longer than the
lungs
Bones: metabolic acidosis alters bone cell function; there is an increase in osteoclastic
bone resorption and a decrease in osteoblastic bone formation. As we age, metabolic
acidosis leads to greater loss of bone mineral and more prone to fracture.
Respiratory acidosis, caused by an increase in the partial pressure of carbon dioxide
induces far less bone dissolution and resorption and the additional hydrogen ions are
not buffered by bone.
15. The main types of disorders of acid-base balance - respiratory acidosis and alkalosis,
metabolic acidosis and alkalosis, their causes. Changes of the basic indicators of acidbase status in acidosis and alkalosis.
Metabolic acidosis causes are
- Increased acid production (diabetes, ethanol poisoning)
- Acid ingestion
- Decreased renal acid excretion (renal tubular acidosis, chronic renal insufficiency
(decreased renal fit and reabsorption)
- GI or renal HCO3- loss
Metabolic alkalosis causes are:
- Acid loss (severe vomiting)
- HCO3 retention
Respiratory acidosis is Pco2> 45 mm Hg (hyperacapnia), causes are:.
- Decrease in minute ventilation (hypoventilation), pneumonia, bronchial asthma
attack,
- Chronic bronchitis
Respiratory alkalosis is PCO2 <35 mm Hg (hyporecapnia), cause are:
- Increase in minute ventilation (hypеrventilation) traumatic brain injury leading to
stimulation of respiratory center
Changes in basic indicators of acid-base status
16. System of hemostasis: role, components. Role of endothelium in hemostasis.
Anticoagulant properties of intact endothelium. Pro-coagulant properties of activated
endothelium and subendothelial layer.
Haemostasis is a process that prevent and stop the bleeding, meaning to keep blood
within a damaged blood vessel (the opposite of hemostasis is hemorrhage). The
hemostatic system comprise 3 processes: platelet aggregation, platelet coagulation and
fibrinolysis. They also termed as primary, secondary and tertiary hemostasis.
Primary hemostasis: vasoconstriction and platelet aggregation
- Components: endothelial cells, platelets, adhesive proteins (vWF, collagen), thrombin
- Role: formation of platelet plug in response to vascular injury to limit the bleeding
Secondary hemostasis: coagulation, polymerase of fibrinogen to fibrin
- Components: cells, enzymatic and non-enzymatic coagulation factors,
phosphatidylserine and Ca2+
- Role: reinforce of the plug by formation of fibrin clot.
- 2 pathways: extrinsic and intrinsic
Tertiary hemostasis: fibrinolysis: lysis of fibrin clot after the injury healed.
- Components: tissue plasminogen activator, plasminogen,
- Role: removal of fibrin clot when it is no longer needed to maintain normal blood flow
of vessel
Role of endothelium in hemostasis:
Anticoagulant properties of intact endothelium include
- Provision of a cell surface with heparin-like molecules (which can serve as binding sites
for antithrombin III to inactivate thrombin and FX
- Thrombomodulin alters the substrate specificity of thrombin by binding to activated
protein C (protein C is activated by binding to protein S) then inhibit thrombin factors: V
and VIII
- Maintenance of a low level of tissue factor and generation of prostacyclin.
The inactivation of thrombin is accelerated by heparan sulfate present on the
endothelial cell surface. Thus, the intact endothelium has the capability of modifying
thrombin action and inhibiting platelet aggregation
Procoagulant properties
- Propagate factor X and prothrombin activation once factor IXa and factor Xa have been
formed.
- Produce and secrete binding site of factors regulating the balance of plasminogen
activator and inhibitor synthesis
Subendothelium is the conective tissue between the endothelium and the inner elastic
membrane in the intima of arteries. Subendothelium consist of collagen, elastic tissues,
proteoglycans and non-collagenous glycoproteins (fibronectin, vWF)
Exposure of this layer after damage of vessel wall is responsible for platelet adherence.
vWF bind with collagen, vWF then undergoes a conformational change and platelets are
capture via their surface membrane glycoprotein GpIB binding to vWF
Tissue factor (TF) is a transmembrane glycoprotein that functions as the primary cellular
initiator of blood coagulation. Perivascular cells express TF and provide a hemostatic
barrier to limit hemorrhage after vessel injury.
Collagen is ideally situated to initiate hemostasis. Collagen is the only matrix protein
which supports both platelet adhesion and complete activation. When collagen
becomes exposed to flowing blood, platelets rapidly adhere, spread, become activated
and begin to form an aggregate.
17. Role of platelets in the blood clotting process. Platelet receptors (GPIIbIIIa, GPIb),
their ligands and functions. Vascular-platelet hemostasis. Stages. The mechanism of
platelet activation.
Role of platelet in blood clotting
Platelets play a major role in blood clotting. When blood vessel is injured, people started
to bleed. The platelets will clot (clump together) to plug the hole in the blood vessel and
stop the bleeding.
Platelets contribute their hemostatic capacity via adhesion, activation and aggregation,
which are triggered upon tissue injury, and these actions stimulate the coagulation
factors and other mediators to achieve hemostasis.
Platelet receptors
Glycoprotein
GPIb
Functions
Adhesion to subendothelium
(vWF)
Process
Binding to vWF immobilized on
collagen becomes adhesion at
the site of damage
Platelet undergoes dramatic
changes to an irregular sphere
with many podia spreading on
subendothelium.
Ligands
vWF
GPIIbIIIa (integrin αIIbβ3)
Aggregation (binding of platelet
to the fibrinogen)
Adherent platelets secrete the
content ADP, thromboxane A2,
thrombin. They stimulate
GPIIbIIIa from low affinity to high
affinity state for binding
fibrinogen and vWB factors.
Fibrinogen acts as bride on
another adjacent activated
platelets, forming aggregation.
vWF, fibrinogen
Vascular- platelet hemostasis
Adhesion- Platelet should be attached to the damaged area, can attach to
subendothelium collagen directly or by means of von Willebrand Factor, there are
different type of receptors they are glycoproteins (GP).
GP Ia/IIa binds platelets to subendothelium collagen directly.
GPIb binds to von Willebrand factors which helps platelets bind more efficiently to
collagen.
Activation- Adhesion of platelets to subendothelial collagen directly or via von
Willebrand factor. Interaction with sub endothelium collagen platelets change their
shape from flat discs to spheres with podia, because of calcium signaling inside the
platelets and adhere to collagen, components of platelets can be secreted, they are
different types of granules in platelets, and they interact with each other they form
aggregates.
There is release of TXa2 involved in activation of platelets, ADP released and can involve
in activation of platelets. Formation of thrombin is the most important activator of
platelets and their interaction happens through GPIIb-IIIa receptors from fibrinogen
molecules
Aggregation- Aggregation occurs due to receptors GPIIb/IIIa (not active on the platelet
as fibrinogen may bind), only after activation the binding may happen.
Secretion-dense granules contain ADP, alpha granules like thromboxane A2 (TXa2) may
activate nearby platelets.
18. The secondary hemostasis. Plasma coagulation proteins. General characteristics.
Thrombin formation. The functions of thrombin. The conversion of fibrinogen to
insoluble fibrin. The role of thrombin and factor XIII.
Plasma coagulation proteins:
Fibrinogen (FI)
- The conversion of fibrinogen to fibrin occurs by cleaving of Arg-Gly peptide bonds of
fibrinogen.
- Fibrinogen has a molecular weight of 3,40,000 D and is synthesized by the liver.
- Normal fibrinogen level in blood is 200-400 mg/dl.
- The fibrin monomers formed are insoluble.
- They align themselves lengthwise, aggregate and precipitate to form the clot.
- Fibrinogen is an acute phase protein.
Serine proteases
- Prothrombin (F II) is a vitamin K-dependent serine protease, which is enzymatically
cleaved to thrombin by activated factor X (FXa). It also activates factors Va, Ca2+,
- Factor VII is vitamin K-dependent serine protease. It initiates coagulation by activating
factors IX and X simultaneously with tissue factor in the extrinsic pathway.
- Factor IX is also known as Christmas factor. It is a proenzyme serine protease, which in
the presence of calcium activates factor X. Its deficiency cause hemophilia B
- Factor X, also known as Stuart-Prower factor, is a serine protease of the coagulation
cascade. In the presence of calcium and phospholipid, FⅩ functions in both intrinsic
and extrinsic pathway of blood coagulation. FⅩ is activated by factors FIX and F VII.
- Factor XI: the zymogen form of factor XIa, one of the enzymes of the coagulation
cascade in intrinsic pathway (cleavage of FX to FXa)
- Factor XII: also known as Hageman factor, its zymogen form of XIIa, function in intrinsic
pathway for cleavage of FXI.
Cofactors:
- V: synthesis occurs in the liver. The molecule of factor V circulates in plasma as single
chain. It is able to bind to activated platelets and is activated by thrombin (requires
Ca2+). The activated factor V (Va) is a cofactor of prothrombinase complex and it
converts prothrombin to thrombin on cell surface membrane
- VIII: During injury, factor VIII is activated and separated from vWF. Factor VIIIa acts as
cofactor for IXa
Transglutaminase XIII - Promotes the cell adhesion through α9β1 or α4β1 intergrins
Formation of thrombin. Functions of thrombin.
- Prothrombinase complex catalyzes the conversion of prothrombin (factor II) which is
an inactive zymogen to thrombin (IIa), an active serine protase
- Thrombin functions
+ Catalyzes the cleavage of fibrinogen and liberation of fibrinopeptide
+ Activators of factors: V, VIII, XI and XIII
+ Induction of platelet aggregation, platelet secretion and platelet procoagulant activity
+ Release of ADP from platelets
The conversion of fibrinogen to insoluble fibrin. The role of thrombin and factor XIII.
Conversion of fibrinogen to insoluble fibrin
- During damage of vessel, platelet plug is formed. These platelets have thrombin
receptors on the surface and they bind to thrombin molecules
- Thrombin converts the fibrinogen which is already in the plug into fibrin, liberates
fibrinopeptides
- Under influence of factor XIIIa, fibrin molecules combine, forming a long fibrin that
entangle platelets, building a spongy mass that hardens and contracts to form the blood
clot.
The role of thrombin and factor XIII
- Thrombin catalyze the cleavage of fibrinogen and then liberate fibrinopeptide
- Factor XIII completes the cross linkage of fibrin so its hardens and contracts. The cross
linked fibrin forms the complete clot
19. The cell model of blood coagulation, the basic processes occurring at each stage.
Stages: initiation, amplification, propagation (formation of fibrin).
The cell model of blood coagulation: the activation of both extrinsic and intrinsic
pathway to produce enough thrombin for blood clotting
Initiation: activation of extrinsic pathway:
- Tissue factor (III) is exposed, forming complex with factor 7 => 7a
- 7a activates f10, then activates f2 (thrombin) but small amount of thrombin is not
sufficient
Amplification: activation of intrinsic pathway
- Thrombin binds to platelets by receptor vWF
- Thrombin activates f11, 8, 5 (which are factors of intrinsic pathway)
- f 11 activates f9m f9a forms complex with f8a together activate f10, forming more
thrombin
Propagation: large amount of p10 is activated
- f10 binds to f5, which is activated by thrombin, results increasing in factor 10 activity
- More thrombin is formed from f2
Fibrin formation:
1. Fibrinogen is cleaved to fibrin (activated factor 1)
2. Fibrin molecules form fibrin strands
3. Thrombin also activates factor 13, which crosslinks individual fibrin into stable
fibrin network
4. RBC’s are also caught in the fibrin network
20. Vitamin K-dependent coagulation factors. The physiological role of γ-carboxylation.
Warfarin, mechanism of action, the main side effects. Vitamin K, food sources, daily
requirement, biochemical functions, causes and symptoms of deficiency.
Vitamin K dependent proteins
Coagulation: factors II, VII, IX, X and Anticoagulant: protein C, protein S.
Factors II, VII, IX and X are required for blood clotting. Protein C and S work together to
regulate and control blood clot formation by inactivating coagulation factor (V and VIII).
The physiological role of γ-carboxylation.
Vitamin K dependent proteins all contain glutamic acid residues. These factors need to
be carboxylated to perform proper function. γ- glutamyl carboxylase is enzyme catalyze
for this carboxylation. CO2 and O2 also required.
Vitamin K is the cofactor for γ- glutamyl carboxylase. In reduced-hydroquinone form, it
works as cofactor. During this reaction, vitamin K is oxidized to epoxide form which is
inactive.
The carboxylation provides negative charge, enable glutamic residues to bind to
Ca2+and thereby to membrane phospholipid of platelets. It then allows interaction with
other molecules to form coagulation
To reactive vitamin K, we need vitamin K reductase with the help of NADH to convert
vitamin K to active reduced form.
- Vitamin k (naphthoquinone) received in the liver is restored by NADPH – dependent
Vit K reductase to form dihydroxyqionone of vitamin K.
- During enzymatic reaction, dihydroxyqionone is oxidised and inactive 2,3 epoxide of
Vit k is formed.
- Epoxide is regenerated into dihydroxyquinone by reductase, which is protein
thioredoxin
Warfarin – Inhibits the synthesis of active forms of Vit k dependent clotting factors
II,VII,IX and X. Warfarin is used as anticoagulant and is used to treat blood clots such as
deep vein thrombosis and embolism to prevent stroke in people.
Side effects of Warfarin :
• Gum bleeding
• Bruises
•
•
•
•
•
•
•
•
•
Severe headache
Blood in urine
vomiting
confusion
deep vein thrombosis
myocardial infarction
Diarrhea
Anemia
Jaundice
Vitamin K is fat soluble vitamin
- K1: phylloquinone (in green vegetable)
- K2: menaquinone (intestinal bacteria)
- K3: synthetic menadione
Food sources: greenly leafy vegetable: spinach, broccoli, lettuce, soybeans and canola
oil
Daily requirement: 80 µg
Biochemical functions:
- Maintenance of normal level of the blood clotting protein factors II, VII, IX, X, protein C
and S which are synthesized in the liver as inactive precursor proteins
+ Confers calcium binding and lipid binding on proteins (II, VII, IX, X)
+ Post-translational modification
+ Converts 1st 7-12 glutamic acid to g-carboxyglutamic acid
+ Without vitamin K, secretes des-g-carboxyglutamic acid containing proteins (inactive
in coagulation)
Causes of deficiency: hemorrhagic syndrome
- Insufficient dietary intake, inadequate absorption, decrease storage of the vitamin due
to liver disease, decrease in intestine production
Eg: malabsorption, celiac disease, chronic pancreatitis, Crohns disease, cystic fibrosis
Symptoms:
- Bleeding of GI tract (due to absent or deficient blood-clotting elements)
- Blood in urine or stool (blood from internal cannot clot and excreted with urine, stool)
- Excessive bleeding from wounds, punctures, injections or surgical sites (due to absent
or deficient blood-clotting elements)
- Heavy menstruation in women (blood cannot clot)
- Easy bruising (due to absent or deficient blood-clotting elements)
- Increased prothrombin time
- Vitamin K deficiency in newborns associated with bleeding within the skull
Diagnosis:
- Prothrombin time (PT/INR) test: investigate the time of bleeding
- Measurement level of vitamin K (only in reference laboratory and results take several
days)
Treatment:
- Vitamin supplement
- Injections
21. Anticoagulants: characterization, function and role: tissue factor pathway inhibitor
(TFPI), antithrombin III, heparin, protein C and S. The system of fibrinolysis. The main
components of fibrinolytic system (plasminogen, tissue plasminogen activator,
urokinase). The main physiological mechanism of activation. Degradation of fibrin by
plasmin, degradation products.
Anticoagulants
a. Tissue factor pathway inhibitor (TFPI).
Tissue factor initiates extrinsic pathway. TFPI is an anticoagulant protein that inhibits
early phase of procoagulant response.
TFPI inhibits FVII-TF complex and FXa
b. Antithrombin III. Role of heparin
Antithrombin III inhibits factor X and thrombin
Heparin: from basophils and mast cell potentiates effects on antithrombin III (together
inhibit IX, I, XI, XII and thrombin)
- Heparin exerts parts of its anticoagulant activity through interaction with
antithrombin. Antithrombin binds specifically to heparin and induced a conformational
change in the antithrombin, accelerate enzyme inhibition to intrinsic pathway cloting
factor (12,11,9,9a,10a, thrombin,)
c. Proteins C and S
- Binding to thrombomodulin to inactivate factor V and VIII
The system of fibrinolysis Fibrinolysis system is activated to lysis the fibrin in blood clot
when there is no damage in epithelial cells anymore.
Sequences:
- Fibrinogen is a dimer which has a kringle-dependent binding site for plasminogen and
binding site for tPA via finger domain
- Plasminogen binds to fibrinogen (different plasminogen have different affinity to
fibrinogen)
- tPA binds to fibrinogen and generate formation of plasmin.
- Plasmin degrades fibrin, forming soluble fragments (fibrin degradation products)
Fibrinolytic factors
Plasminogen: inactive proenzyme form of plasmin.
- It is the major enzyme that degrades fibrin clots and circulates in the blood.
- 2 types of plasminogen (based on terminal end): glu-plasminogen and lys-plasminogen
- It is inhibited by a2 antiplasmin
- Plasminogen can bind to both fibrin and fibrinogen. Inactive tPA (tissue plasmin
activator) is released from vascular endothelial cells due to injury. Then it binds to fibrin
and be activated. Active tPA converts plasminogen to plasmin.
Tissue plasminogen activator: endogenous activator of plasminogen in the blood
- Single-chain molecule (68kDa glycoprotein) in vascular endothelial cells and secreted
into plasma under stimulation. Binding to fibrin concentrates and orientates tPA and
plasminogen, inducing conformational changes that promote efficient clot lysis.
- Rapid fluctuation of tPA concentration is observed in response to exercise, venous
occlusion, alcohol, drugs
- Inhibitors: PAI-1 (plasminogen activator inhibitor-1).
- tPA cleavages plasmin and produces more active two-chain molecule.
Urokinase
- Mainly produced in the kidneys as inactive single chain molecule (scu-PA).
- Function in tissue-related proteolysis and play a secondary role to tPA as physiological
activator in the blood
- Activation of inactive single chain molecule (scu-PA) results increase plasmin
- Does not bind to fibrin and is not activated by fibrin
Degradation of fibrin under influence of plasmin
- After plasminogen cleavage, plasmin is formed. It will degrade fibrin or fibrinogen
resulting soluble fibrin degradation products
MODULE VIII. BIOCHEMISTRY OF TISSUES
1. Metabolism of the kidneys. Features and differences of metabolism in renal cortex
and in renal medulla. Aerobic and anaerobic oxidation processes, their localization in
the kidney. Gluconeogenesis. Role of kidneys in the synthesis of biologically active
substances (creatine, erythropoietin, 1,25-dihydroxycholecalciferol). The processes of
formation of urine: filtration, reabsorption and secretion.
The kidney can be considered two organs due to differences in the distribution of
various enzymes in the renal cortex and medulla.
Renal cortex:
Glucose synthesis
• Cells of cortex have considerable amounts of gluconeogenic enzymes (but little
hexokinase): G6P, Pyruvate carboxylase, PEP carboxylase
• Most important substrate for gluconeogenesis is glutamine, lactate, and
glycerol.
• The release of glucose by the normal kidney is exclusively, a result of renal
cortical gluconeogenesis.
Renal medulla:
Glucose utilization
• Cells of medulla have considerable amount of hexokinase (of glycolysis). They
can take up phosphorylate and metabolize glucose through glycolysis (but don’t
have gluconeogenic enzymes)
• Glycolytic enzymes: Hexokinase, phosphofructokinase, Pyruvate kinase
• Glucose can be used for reabsorption of urine, released to the bloodstream to
maintain blood glucose level.
Kidney needs very intense energy metabolism
Aerobic process (produce more energy): Tubular epithelial cells in renal cortex, have
better blood supply and more mitochondria, place for aerobic processes (produce
more energy).
These cells can utilize almost all nutrients: glucose, fatty acids, ketone bodies and amino
acids (significance of glutamine)
Anaerobic Process Metabolism of cells in renal medulla is limited by an insufficient O2
supply, most ATP molecules is thus obtained in anaerobic glycolysis.
Kidneys represent a significant site of gluconeogenesis (pathway used by body to create
glucose), reverse to glycolysis (only diff in 3 reactions)
- Free amino acids (glutamine, proline), lactate, pyruvate, fructose, fatty acids, ketone
bodies are substrates for gluconeogenesis forming G-6-P then glucose.
+ Lactate: (from Cori cycle) waste product of muscles, travel through the blood stream
+ Free aa: from absorption, broken down to oxaloacetate (by deamination) and release
NH3+
+ Oxidation of free fatty acids provide stimulation to gluconeogenesis (activates
enzymes, provide energy)
- Glucose which is produced via gluconeogenesis can provide energy for urine formation
and maintain blood glucose level require for brain function.
Synthesis of biologically active substances
• Creatine: In males- 1.5 to 2.0g/24hr (7.1-17.7 mmol/day)
In female: 0.8 to1.5g/24hr (5.3-15.9 mmol/day)
Creatine is the end product of nitrogen metabolism. It is found in the muscle tissue
from creatine phosphate.
Creatine enters urine mainly by glomerular filtration and in an extremely small amount
due to active tubular secretion
• Erythropoietin: Stimulates RBCs synthesis, produced by kidneys under hypoxia
condition or glucocorticoid influence.
- Glucocorticoid increases Na+ absorption in kidney -> increase O2 utilization ->
hypoxia
A) decrease in O2 delivery to kidneys (hypoxia)
B) Peritubular intestinal cells detect low O2 in blood and secrete EPO in blood
C)Proerythroblasts in red bone marrow mature quickly into reticulocytes
D)More reticulocytes enter circulating blood
E) Large no. of RBCs in circulation
F) Increased O2 delivery to tissues
G) return to homeostasis when response brings O2 back to normal
• 1,25 dihydroxycholecalciferol:
Active form of vit D3 (calcitriol)
Production of Vit D is controlled by PTH and serum phosphate concentration: a rise in
PTH or fall in serum phosphate increases 1,25 dihydroxycholecalciferol synthesis which
then increase Ca2+ absorption in kidney, bone, and intestine.
The processes of formation of urine: filtration, reabsorption, and secretion.
Filtration: Exit to the Bowman’s capsule of everything that is in the blood, except for the
cells and proteins. It is done using the high blood pressure that occurs in the kidney
Moves: water, urea, uric acid, glucose, amino acids, salt
Almost 180l per day. Most filtrated is reabsorbed, leaving about 1L of fluid to be
excreted each day. Can be measured by GFR
Reabsorption: Reverse absorption of renal tubules of necessary substances (ions,
glucose, aa)
- Occurs mostly in PCT (ions, glucose, aa, water) and less in DCT (water, urea)
Function
- Moving necessary molecules back to the blood from the nephron.
- Concentrate the urine and accumulate urea.
Moves: Osmosis (moves water)
Active (using ATP)- glucose, Aa, Na+, K+,H+, ClDiffusion – urea
GFR = 180l/day
UFR= 1.5l/day
- In the loop of Henle - some NaCl moves back to blood.
Excretion: It is active or passive transition of compounds from epithelial cells to primary
urine
- Late distal convoluted tubule
-
Function: molecules move from blood to nephron
Moves: uric acid, ammonia, H+ ions
Collecting duct
Function: urine is formed and travels to the ureter
Moves: water, salt, urea, uric acid
2. Water balance. The role of the skin, lungs, gastrointestinal tract and kidneys in
removing water. Factors of water balance in the body - blood osmolality, volume of
blood, blood pressure, concentration of sodium and potassium. The regulation of water
reabsorption. The role of antidiuretic hormone. Hypoproduction of antidiuretic
hormone, manifestations.
Water balance: balance between total amount of water entering the body through
ingestions of liquid and food and the total output of water by kidneys, lungs, skin
Role of skin, lungs, GI tract and kidney in removing of water:
- Skin: sweat glands release excess water and salts
- Lungs: exhale carbon dioxide and water vapor
- Kidneys: filter and clean the blood to produce urine
- GI tract: (main site of absorption of water for distribute throughout the whole body)
small part of water is excreted with feces.
Factors of water balance in the body
- Blood osmolarity (caused by albumin): thirst center is stimulated by an increase of blood
osmolarity, lead to increase intake of water
- Volume of blood: low volume of water in blood will increase blood osmolarity and then
stimulate thirst.
- Blood pressure: in response to low blood pressure, kidney release antidiuretic hormone
which acts on kidney to increase water reabsorption then increase blood volume and then
blood pressure
- Concentration of sodium and potassium: regulate extracellular fluid. Water follows the
sodium due to osmosis. Potassium has little effect on osmotic pressure but due to
sodium-potassium pump it helps to regulate flux of sodium.
Regulation of water reabsorption:
- Hormones: aldosterone, AHD
+ AHD is produced by hypothalamus when osmolarity of plasma rise or due to high sodium
concentration detection by osmoreceptors, it triggers the water channel in collecting
ducts of the kidneys to reabsorb water. Decreasing of osmolarity inhibits secretion of AHD
+ Aldosterone: is produced in zona glomerulosa of adrenal cortex, regulates Na+ K+
exchange and Na+ H+ exchange at renal tubules
- Renin-angiotensin system: when ECF volume fall due to decreased blood pressure, salt
depletion, prostaglandins, renin is secreted by kidney. It stimulates forming of angiotensin
II which acts as vasoconstrictor, increase aldosterone biosynthesis for renal action and
sympathetic nervous stimulation (for secretion of ADH in hypothalamus)
Role of antidiuretic hormone: control blood pressure by acting on kidney to regulate
water level. It conserves fluid volume of the body by reduce amount of water passed out
to urine.
Hypoproduction of antidiuretic hormone causes diabetes insipidus
- The hypoproduction or insensitivity of ADH causes an inability to concentrate urine in
distal renal tubules resulted dilute urine. The person with this condition passed urine with
low osmolarity.
- Causes: damage (tumor, trauma) of hypothalamus or posterior pituitary gland, primary
polydipsia (excessive water drinking)
- Manifestations: increasing amount of dilute urine, dehydration and low blood pressure.
- Diagnosis: ADH blood test, urine test, plasma osmolarity and sodium level
3. The regulation of sodium reabsorption. Renin-angiotensin-aldosterone system (RAAS).
The scheme, role of the RAAS in sodium reabsorption. The mechanism of hypertension in
hypoperfusion of kidneys.
Under dropped blood volume and dropped blood pressure conditions, sodium
reabsorption is stimulated by renin-angiotensin-aldosterone system. The main
purpose of RAAS is increase blood volume and blood pressure. RAAS involves 3
main roles of kidney, liver, and lungs.
1st Role (By kidney -> release of RENIN) in response to two stimuli
Kidney must maintain the glomerular filtration rate. So if blood volume drops (due to
haemorrhage or peripheral vasodilation in case of anaphylactic shock), blood pressure
also drop, GFR decreases, filtration rate of Na+ decreases, less Na+ can reach DCT.
Receptors sense the change in pressure and osmolarity, kidney secretes renin from
juxtamedullary.
2nd Role (By liver -> synthesis of Angiotensinogen)
Angiotensinogen is synthesized in liver and released to blood stream, where it is
activated by Renin to ANGIOTENSIN I
3rd Role (By LUNG -> production of ANGIOTENSIN CONVERTING ENZYME ACE
Angiotensin converting enzyme ACE exists in kidney, but mainly it is produced in the
lungs. ACE converts angiotensin I to angiotensin II, which plays important roles
1- Systemic vasoconstriction, so increases BP
2- Constricts the efferent arteriole leading to backing up of blood into the glomeruli,
which in turn increases the glomerular filtration rate GFR and increase the Na⁺
reabsorption in distal convoluted tubules
3- Stimulate the release of anti-diuretic hormone ADH from the posterior pituitary
gland, which results in increase water reabsorption from distal tubules and
collecting ducts, so increase blood volume and increase blood pressure.
4- Also, it acts on hypothalamus to stimulate water intake the sense of thirst 5Stimulate release of ALDOSTERON from the renal cortex
Mechanism of hypertension in hypoperfusion of kidneys:
- High blood pressure filtrates sodium through glomerular to GFR and sodium level
influence the reabsorption of kidney.
- Hypoperfusion (which caused by low blood pressure) of kidneys decreases sodium
level in GFR.
- Arterioles constrict to increase pressure which then increase filtration, increase sodium
level in GFR and cause hypertension
4. The role of the kidneys in maintaining the acid-base status – reabsorption of
bicarbonate, secretion of H+, ammonium, excretion of organic acids.
Kidney can regulate the pH of the extracellular fluid by excrete or reabsorb H+ ion.
Normal urine has a pH around 6. pH of the urine may vary from as low as 4.5 to 9.8,
depending on the amount of acid excreted.
The major mechanisms for pH regulation of kidney are:
Secretion of H+
Reabsorption of bicarbonate (recovery of bicarbonate)
Excretion of titratable acid (net acid excretion)
Excretion of NH4+ (ammonium ions).
Excretion of H+; reabsorption of bicarbonate
- This process occurs in the proximal convoluted tubules, distal tubule and loop of
Henle
- The CO2 combines with water to form carbonic acid, with the help of extracellular
carbonic anhydrase. The H2CO3 then ionizes to H+ and bicarbonate.
- The hydrogen ions are secreted into the tubular lumen in exchange for Na+
reabsorbed. Na+ ions forming salt with HCO3 – is reabsorbed into the blood.
- H+ which is secreted to the lumen under catalyze of intracellular carbonic anhydrase,
create CO2 which diffuse through membrane and get to tubular cells. Then undergoes
ionized again.
- There is net excretion of hydrogen ions, and net generation of bicarbonate. So this
mechanism serves to increase the alkali reserve.
Excretion of H⁺ as Titratable Acid
- Phosphate in plasma present in 2 forms: monophosphate ion, diphosphate ion.
- Under influence of intracellular carbonic anhydrase, ion H+ is formed from intracellular
CO2
- H+ ions are excreted through hydrogen or hydrogen-potassium channels, bind with
monophosphate ion, forming titratable acid (diphosphate acid) for excretion. HCO3- is
reabsorbed to the blood in basolateral surface.
Excretion of Ammonium Ions
- Ammonia (NH3) is synthesized from glutamine (by deamination)
+ In PCT, ammonia is synthesized by tubular cells, then diffuse to the lumen
+ In collecting duct, ammonia diffuses from medullary interstitum (outside tubular cells)
into the lumen
- H+ ion which is excreted to the lumen through H+ channel or H+/K+ channel (product
of intracellular carbonic anhydrase reaction) creates NH4+ ion for excretion
5. General properties of urine: quantity, color, clarity, relative density, pH. Changes in
pathological situations. Organic and inorganic components of urine. Pathological
components - protein, glucose, bile pigments, ketones, erythrocytes, enzymes.
General properties of urine:
Quantity: average urine production in human is about 1-2 L per day.
Chemical composition of urine: Urine is an aqueous solution of greater than 95% water,
with a minimum of these remaining constituents, in order of decreasing concentration:
- Urea 9.3 g/L
- Chloride 1.87 g/L
- Sodium 1.17 g/L
- Potassium 0.75 g/L
- Creatinine 0.67 g/L
- Other dissolved ions, inorganic and organic compounds (proteins, hormones,
metabolites)
Color: typical yellow-amber but varies according to recent diet and the concentration
of the urine.
Dilute urine has light yellow color. Dark urine (concentrated) may indicate dehydration.
Red urine indicates red blood cells within the urine, a sign of kidney damage and disease.
Tubrbidity (clarity): clear, slightly cloudy, cloudy, opaque, or flocculent.
Excess turbidity results from the presence of suspended particles in the urine. Common
causes of abnormal turbidity include: increased cells, urinary tract infections or
obstructions.
The pH of normal urine: is generally in the range pH 4.6 – 8, with a typical average being
around 6.0
Density: (ratio of the weight of a volume of a substance compared with the weight of
the same volume of distilled water). The density of normal urine ranges from 0.001 to
0.035
Changes in pathological situations:
- Proteinuria: protein content in urine, often due to leaky or damaged glomeruli
- Oliguria: an abnormally small amount of urine, often due to shock or kidney
damage
- Polyuria: an abnormally large amount of urine, often caused by diabetes
- Dysuria: painful or uncomfortable urination, often from urinary tract infections
- Hematuria: red blood cells in urine, from infection or injury
- Glycosuria: glucose in urine, due to excess plasma glucose in diabetes, beyond
the amount able to be reabsorbed in the proximal convoluted tubule.
Organic and inorganic components of urine:
The total amount of solids in a 24-hour urine sample averages around 60g: 35g are
organic and 25 g are inorganic.
- Inorganic components: Chloride, Sulphate, Calcium, Inorganic phosphate, Ammonia.
Sodium Chloride is the predominant Chloride and makes up about half of the inorganic
substances.
- Organic components: Urea, Uric Acid, Creatinine, Ethereal Sulphate (Organic
Sulphate), urobilinogen.
Pathological components : normally not present in urine or in too high/low level
Proteins :
- With excess of proteins => proteinuria (glomerulonephrititis, bacterial infection,
pregnancy)
- 150mg pf proteins are excreted daily
Glucose:
- With excess of glucose => glycosuria (diabetes mellitus, emotion stress)
- Renal threshold for glucose is around 10 mmol/L
Ketone:
- Excess level of ketone bodies => ketonuria (starvation, diabetes, pregnancy)
Erythrocytes:
- In the absence of contamination, the presence of more than 3 to 5 erythrocytes
per field on more than one occasion may signal one of several kidney or urinary
tract infections
Bile pigments are due to catabolism of the hemoglobin are:
- Bilirubin is orange or yellow in color
- Biliverdin is green in color
-
Urobilinogen.
6. Muscle proteins. The structure of the myofibrils and myofibrillar proteins. Structure
and properties of myosin. The enzymatic activity of myosin. Thin (actin) filaments,
structure, composition. Structure of the thick filament.
Muscle proteins: They composed of actin, myosin and actin crosslinking proteins,
tropomyosin and troponin. Others: myoglobin, collagen, enzymes etc.
Myofibrils- muscle fiber unit.
- Myofibrils are aligned and give distinct bands [I band (light)and A band(dark)]
- Composition: They contain 2 different types of filaments
+ Thick- myosin (15nm) in A band
+ Thin – actin (6nm) in I band and located at periphery of A band
I- band: light band contains only thin filaments and Z disc is dark area of I-band
A- band: dark band contains entire length of thick filaments and 1 part of thin filaments
-H zone- light area of A band (bare zone): no sliding
-M line: center of H-zone contains tiny protein rods that hold adjacent filaments together
Properties: Myofibril has ability to contract and relax by sliding mechanism
Myofibrillar proteins: myosin, actin, regulatory proteins (tropomyosin and myosin)
MYOSIN :(PREDOMINANTLY MYOSIN II)
• Composed of six polypeptide chains (hexamer)
• Contains 1 pair of heavy chain and 2 pair of light chain
• Head has a binding site for actin and ATP
• Most of the heavy chain myosin has Alpha helical structure in which 2 chains coil
around each other to form “tail” of myosin
• 4 light chains and N terminus of each heavy chain form 2 globular “heads” on
myosin molecule
Enzymatic activity of myosin – muscle relaxation (sliding filament theory): transform
ATP to ADP+ Pi
- During relaxation phase of the muscle contraction, the head of myosin hydrolyses ATP
to ADP + Pi. This results in formation of high energy ADP-Pi myosin complex
- On contraction, the muscle gets stimulated (through participation of actin, Ca2+,
troponin, tropomyosin) to finally form actin- myosin-ADP-Pi complex
- Next is the power stroke which drives movement of actin filaments over myosin
filaments, followed by the release of ADP and Pi and a conformation change in myosin.
The actin-myosin complex is in a low energy state
- A fresh molecule of ATP now binds to form actin- myosin ATP complex
- Actin, released as myosin – ATP has low affinity for actin. This step is crucial for
relaxation which is dependent on binding of ATP to actin- myosin complex.
A fresh cycle of muscle contraction and relaxation now commences with hydrolysis of
ATP and the formation of ADP-Pi-myosin complex
-Ultimately ATP is immediate source if energy for muscle contraction
Sources of ATP:
• By substrate level phosphorylation of glycolysis using glucose or glycogen
• By oxidative phosphorylation
• From creatine phosphate
• From 2 molecules of ADP in a non-catalyzed by adenylyl cyclase
ACTIN filament:
• Major constituent of thin filaments of sarcomere
• Exists in 2 forms- monomeric G-actin (glomerular) and polymeric F- actin filament
• G-actin constitutes of about 25 % of muscle proteins by weight
• In presence of Mg2+, G-actin polymerizes (non-covalently) to form an insoluble
double helical F-actin with thickness 6-7 nm
• Cross- linking proteins found in association with actin are: Tropomyosin and
Troponin
• Tropomyosin, composed of two chains attaches to F-actin in the grooves.
• Troponin consists of 3 polypeptide chains- Troponin (T-binds to tropomyosin, Ifor inhibition, C- for Ca2+
7. Features, stage and the chemistry of muscle contraction. Function of troponin
subunits. Energy supply for muscle contraction. The regulation of contraction and
relaxation of muscles.
Muscle contraction:
- On contraction, the muscle gets stimulated (through participation of actin, Ca2+,
troponin, tropomyosin) to finally form actin- myosin-ADP-Pi complex
- Next is the power stroke which drives movement of actin filaments over myosin
filaments, followed by the release of ADP and Pi and a conformation change in myosin.
The actin-myosin complex is in a low energy state
- A fresh molecule of ATP now binds to form actin- myosin ATP complex
- Actin, released as myosin – ATP has low affinity for actin. This step is crucial for
relaxation which is dependent on binding of ATP to actin- myosin complex.
A fresh cycle of muscle contraction and relaxation now commences with hydrolysis of
ATP and the formation of ADP-Pi-myosin complex
-Ultimately ATP is immediate source if energy for muscle contraction
Function of troponin subunits:
• Troponin is component of thin filaments
• It is a complex of 3 regulatory proteins is integral to non-smooth muscle
contraction in skeletal as well as cardiac muscle
1)Troponin C: has Ca2+ binding ability. Up to 4 Ca2+ ions can bind per molecule of
troponin C
2)troponin I: inhibitory protein; inhibits F-actin myosin interaction
3)Troponin T: binds to troponin- tropomyosin complex
• Troponin is attached to tropomyosin sitting in the groove between actin
filaments in muscle tissue.
Energy supply for muscle contraction:
• By glycolysis, using blood glucose or muscle glycogen
• By oxidative phosphorylation
• From creatine phosphate
• From 2 molecules of ADP in a reaction catalyzed by adenylyl cyclase
Regulation of Muscle contraction
A) Skeletal: depends on level of Ca2+, tropomyosin, ADP, myosin, actin
- Ca2+ triggers the binding of actin and myosin head for sliding.
- Tropomyosin inhibits binding of actin and myosin head
B) Smooth
- SMC lacks of troponin, but it has calmodulin
- Ca2+ binds with calmodulin and forms complex. Calmodulin/Ca2+ activates myosin
kinase
- Myosin kinase phosphorylates myosin for binding with actin
Relaxation of muscles regulation
A) Skeletal
• Fall in Ca2+ transient
• Sensitivity of troponinC to Ca2+ decreases
• Cross bride dissociation
B) Smooth
SMC relaxation occurs either because of
• Removal of contractile stimulus (neurotransmitter
• By direct action of a substance that stimulates inhibition of the contractile
mechanism
• Na+/Ca2+ exchangers located on the plasma membrane aid in decreasing
intracellular Ca2+.
8. Changes in metabolism during muscular work. Features of metabolism in muscle
tissue. Types of muscles.
Changes in metabolism during muscular work: skeletal muscular work have high energy
requirement, lower requirement in smooth and cardiac muscle
- ATP is required for pumping of Ca2+ from sarcoplasmic reticulum to SR (initiation of
muscle contraction)
- Metabolism provides ATP for muscular relaxation. Binding of ATP helps to release
myosin head from actin (return to relax state)
During both relaxation and initiation of contraction, ATP level decreases.
Muscle stores small amount ATP. To maintain ATP resynthesis, there are 2 pathways:
- Substrate-level phosphorylation (anaerobic)
- Oxidative phosphorylation from carbohydrate and fat metabolism (aerobic)
Types of muscles: skeletal, cardiac, and smooth muscles.
Cardiac muscles:
- Mechanism of contraction is similar to skeletal muscle: Ca2+ bind to troponin C exposes
myosin binding site on actin
- Sources of calcium: intracellular from sarcoplasmic reticulum and extracellular calcium
enters through calcium channels
Smooth muscles:
- Lacks troponin but contraction is still regulated by cytoplasmic calcium levels.
Contraction occurs by binding of calcium to calmodulin
- Sources of calcium: intracellular from sarcoplasmic reticulum and extracellular calcium
enters through calcium channel
9. Features of the biochemistry of the myocardium and the smooth muscle. The
regulation of contractility and relaxation of smooth muscle.
Cardiac muscles:
- Mechanism of contraction is similar to skeletal muscle: Ca2+ bind to troponin C exposes
myosin binding site on actin
- Sources of calcium: intracellular from sarcoplasmic reticulum and extracellular calcium
enters through calcium channels
Smooth muscles:
- Lacks troponin but contraction is still regulated by cytoplasmic calcium levels.
Contraction occurs by binding of calcium to calmodulin
- Sources of calcium: intracellular from sarcoplasmic reticulum and extracellular calcium
enters through calcium channel
Regulation of contraction and relaxation of smooth muscle
- Activators: Release of Ca2+ from sarcoplasmic reticulum: norepinephrine, angiotensin
II, endothelin
- Inhibitors: Removed agonists (mentioned above), vasodilator (atrial natriuretic factor
10. The role of oxygen to the myocardium and metabolic disorder of the heart muscle in
patients with coronary disease. Effects of reactive oxygen species and lipid peroxidation
on the myocardium. Myocardial biochemical changes in patients with coronary heart
disease. Modern markers of heart failure. Markers of acute myocardial infarction.
11. White adipose tissue. Functions. Features of carbohydrate and lipid metabolism in
white adipocytes. Endocrine function of white adipose tissue.
White adipose tissue:
Composition:
- Triacylglycerols: main content that contain saturated and unsaturated fatty acids. This
makes depot fat in a fluid state at body temperature.
- Little phospholipids and cholesterol.
Localization
- Under skin (subcutaneous fat) and in the breast.
- Around Important organs e.g. kidney.
- In the momentum and mesentery.
Sources:
- Digestion
- Carbohydrate storage (by lipogenesis).
Functions: depot fat Is Important for:
- Energy production: During fasting, the triacylglycerol stored in depot fat provide the
body with free fatty acids that are oxidized to give energy.
- Fixation of some organs e.g. kidney.
- Heat insulator around the body.
- Production of vitamin D3: Exposure of skin to ultraviolet rays of sun transforms 7
dehydrocholesterol into vitamin D3.
Adipose tissue uses glucose to form glycerol 3-phosphate (glycolysis), which is essential
for synthesis of TAG. In white adipose tissue two processes control the amount of TAG:
Lipogenesis and lipolysis
Lipogenesis: the synthesis of (TAG) triacylglycerol from Acyl CoA and glycerol (glycerol3-phosphate).
- Activation of fatty acids into fatty acyl
CoA
- In adipose tissue, glycerol phosphate is formed from glucose by glycolysis. Adipocyte
can only uptake glucose under influence of insulin.
Regulation of lipogenesis: Insulin (+) glucagon (-)
1. After meal, lipogenesis is stimulated: Insulin Is secreted which stimulates glycolysis.
Glycolysis supplies dihydroxyacetone phosphate that is converted into glycerol
phosphate in adipose tissue.
2. During fasting lipogenesis is inhibited, as anti-Insulin hormones e.g. glucagon are
secreted. These Inhibit lipogenesis and stimulate lipolysis.
Lypolysis: the hydrolysis of (TAG) triacylglycerols into glycerol and fatty acids. Its
location is the cytosol of adipose tissue cells.
- Lipolysis is carried out by a number of lipase enzymes, which are present in adipose
tissue. These are:
- Hormone sensitive trlacylglycerol lipase. (1st reaction)
- Diacylglycerol lipase. (2nd reaction)
- Monoacylglycerol lipase. (3rd reaction)
Fate of products of lipolysis:
1. Fate of fatty acids:
- Oxidation by tissues to give energy.
- Fatty acids may remain in adipose tissue to be re-esterified into trlacylglycerols
again.
2. Fate of glycerol:
Glycerol may diffuse to blood and then taken up by the liver to give:
- Glucose by gluconeogenesis.
- Pyruvate by glycolysis.
- Triacylglycerols by lipogenesis (outside adipose tissue). Because there is no
glycerokinase in adipocyte which is essential for convert glycerol to glycerol
phosphate for re-esterication of fatty acids
Regulation of lipolysis: The key enzyme controlling lipolysis is the hormone sensitive
triacylglycerol lipase. It exists in 2 forms: active (phosphorylated) and inactive
(dephosphorylated) Insulin (-) glucagon (+)
1. During fasting:
a) Many hormones (epinephrine, norepinephrine, and glucagon) are secreted. They
stimulate adenylate cyclase enzyme
formation of cAMP
Activation of protein
kinase leads to Phosphorylation of hormone sensitive triacylglycerol lipase, which
stimulates lipolysis.
2. After meal:
a) Insulin is secreted
Stimulation of both phosphodiesterase and lipase phosphatase
dephosphorylation and inactivation of hormone sensitive triacylglycerol lipase
àInhibition of lipolysis
b) Caffeine is a substance present in coffee. It inhibits phosphodiesterase enzyme
stimulation of lipolysis.
Endocrine function of white adipose tissue:
Adipose tissue also expresses receptors for most of these factors that are implicated in
the regulation of many processes including food intake, energy expenditure,
metabolism homeostasis, immunity and blood pressure homeostasis
12. Brown and beige adipose tissue. Functions. Features of the metabolism of brown
and beige adipocytes.
Brown fat breaks down blood sugar (glucose) and fat molecules to create heat and help
maintain body temperature. Cold temperature activates brown fat, which leads to
various metabolic changes in the body.
BAT activity in humans is stimulated by cold exposure and by several factors such as
diet and metabolic hormones.
BAT function is regulated at two levels:
- an acute process involving the stimulation of the intrinsic thermogenic activity of
brown adipocytes
- a chronic process of growth involving proliferation or pre-existing brown adipocytes.
BAT is regulated by environmental and nutritional factors that are mediated by neural
and endocrine mechanisms.
13. The proteins of connective tissue: classification, functions. Features of the structure
and function of collagen, ellastin, fibronectin. Stages of formation of collagen fibers. The
role of vitamins and minerals. Scurvy.
Collagen:
- most abundant protein in our body (25-35% of total protein weight)
- produced by fibroblasts
- There are 19 different types of collagen, and they are different in primary structure,
localization, and function.
- Collagen I- found in skin, ligaments, bones
- Collagen II-found mainly in cartilage, vitreous body
- Collagen III-found mainly in skin, muscles
- Collagen IV-found in basal membranes
- Collagen V-found in interstitial tissue
- Collagen VI found in interstitial tissue
- Collagen VII found in epithelial tissues
- Collagen has ability to form insoluble elastic fibers.
Functions:
- Gives strength, support, and shape to the tissues
- Collagen contributes to proper alignment of cells, which in turn helps in cell
proliferation and their differentiation to different tissues and organs
- Collagen (that is exposed in blood vessels) contributes to thrombus formation
Main amino acid component of collagen is glycine. Approximately 1/3 of amino acids
are contributed to glycine. Amino acid sequence of collagen is (Gly-X-Y)n where X and Y
are other amino acid. Collagen contains large quantities of hydroxyl amino acids (proline
and lysine)
Structure:
- All types of collagen are composed of tropocollagen subunits arranged in long axis.
- Each subunit composed of 3 similar polypeptide chains twisted around each other to
form a rod like molecule of 1.4 nm diameter and about 300nm length. There are
forming of hydrogen bonds between polypeptide chains by hydroxyproline residues.
Genetic disorders:
-Ehlers’s Danlos syndrome: a group of inherited disorders characterized by hyper
extensibility of skin and abnormal tissue fragility
-Osteogenesis imperfecta: characterized by abnormal fragility of bones due to decreased
formation of collagen
Elastin:
Structure:
- Elastin is insoluble protein polymer synthesized from a precursor TROPOELASTIN.
- Tropoelastin is a linear polypeptide composed of about 700 amino acid reisidues that
are primarily small and non-polar (eg.- glycine, alanine and valine)
- Tropoelastin is secreted by the cell into the extracellular space. It interacts with
specific glycoprotein microfibrils, such as FIBRILLIN, which functions as a scaffold onto
which tropoelastin is deposited
- High elasticity of elastin is due to crosslinks
Amino acid composition:
- Elastin is rich in glycine, lysine, and alanine residues
Function:
- Ability to stretch under influence of load and restore original size when load is
removed
- Found in large quantities in lungs, arterial blood vessels, elastic ligament, etc.
Williams syndrome: Genetic disease due to impairment in elastin synthesis. The
connective tissue and CNS are affected
Fibronectin
Structure: a high-molecule weight (440kDa) glycoprotein of extracellular matrix. It is
composed of 2 anti-parallel B-sheet linked by disulfide at C-terminal ends.
Each polypeptide chain of fibronectin contains aa sequence: Arg-Gly-Asp
Fibronectin contains 3 modules: type I, II and III.
- Type I and II are stabilized by disulfide bonds
- Type III modules do not contain any disulfide bonds
Function: Fibronectin provide structuring of matrix of CT and cell adhesion due to
interaction with components of extracellular matrix.
There are 2 types of fibronectin
Soluble plasma fibronectin
Insoluble cellular fibronectin
Major component of extracellular matrix.
Location, Major component of blood plasma and
It is secreted by various cells, primarily by
sources produced by the liver
fibroblast
Deposit at the site of wound during later
Function
Strengthen the plug and the clot
stage and promote formation of
granulation tissue
- Fibronectin also plays major role in cell adhesion, growth, migration and differentiation,
wound healing, and embryonic development.
Collagen synthesis:
Stages of collagen synthesis:
- Synthesis of polypeptide chain
- Hydroxylation and glycosylation
- Formation of triple helix: lysine residues undergoes oxidation deamination, catalyzed
by lysyl oxidase forming covalent bond (aldol crosslinking) between lysine residues 2
neighboring polypeptide.
- Elimination from cell into extracellular matrix
- Cleavage procollagen, connect of tropocollagen, forming collagen fiber
Intracellular glycosylation and hydroxylation
Glycosylation: hydroxylysine residues are important for linkage with glycosidic residues
(glucose, galactose, oligosaccharide) by forming O-glycosidic bond between
carbohydrate and hydroxyl group of hydroxylysine
Hydroxylation of post-translation collagen polypeptide occurs in ribosomes (proline
residues)
For synthesis of hydroxyl-derivatives of amino acids, proline is hydroxylated by enzyme
prolyl hydroxylase in the presence of ascorbic acid. Prolyl hydrolase contains Fe2+ in
active center. The presence of ascorbic acid is needed to maintain reduced state of
iron
Extracellular formation of collagen fibers
- Tropocollagen molecules are synthesized by fibroblast in the procollagen form which is
secret into the extracellular matrix.
- Procollagen is converted into tropocollagen by cleavage of C and N-terminal by enzyme
procollagen peptidase
- The connection of tropocollagen forming collagen fiber.
Role of vitamin and minerals
- Act as cofactor for various essential reactions for functional and structural roles in the
body
- Electrolyte circulating
Scurvy resulting from lack of vitamin C ascorbic acid
- Symptoms: weakness, decrease red blood cell, gum disease, skin bleeding, poor wound
healing
- Causes: eating habits, alcohol, intestinal malabsorption
- Treatment: vitamin C supplement
14. Glycosaminoglycans of connective tissue. Features of the structure and function.
Matrix metalloproteinases: classification, functions.
Glycosaminoglycans: long linear polysaccharides consist of repeating disaccharides units
which composed of uronic sugar and amino sugar. They are high polar and attract water.
They are used as lubricant or shock absorber in human body.
Chondroitin
sulfate/dermatan
sulfate
Ketaran sulfate
Synthesized
In Gogi apparatus. Core protein is made in
rough endoplasmic reticulum
In central nervous
system
Localization
Line epithelial
surface, ciliary
epithelium and
plantar epidermis
Skin, blood vessels,
heart valves, tendons
and lungs
Cornea, cartilage,
bone
Structure
Composed of linear
Commonly
polysaccharides
composed of dassembled as
glucuronic acid/ddisaccharide units
glucosamine/lcontaining a
iduronic acid linked
hexosamine, N-acetyl
Ngalactosamine and
acetylglucosamine
glucuronic acid
Linear polymer
consists of
repeating
disaccharide unit
composed of Dgalactose and Nacetyl D
glucosamine 6phosphate
Core proteins
include lumican,
keratocan,
mimecan,
fibromodulin,
aggrecan,
osteoadherin
Non-sulfated
Composed of
linear
polysaccharides
assemble as
disaccharide units
containing
glucuronic acid
and N-acetyl
glucosamine
Function
Regulates biological
activities:
development,
angiogenesis, blood
coagulation, tumor
metastasis
Participates in
development and
glial scar formation
following injury
HA binds cells
together, maintain
shape of eyeballs,
lubricating joints
and surfaces such
as cartilages
Classification
Heparin/Heparan
sulfate
Metalloproteinase:
Role in coagulation,
cardiovascular
disease, infection,
carcinogenesis,
wound repair
Hyalyronic acid
By integral
membrane
syntheases
Major component
of synovial fluid
and tissues,
ground substances
of other
connective tissue
- Collagenase (MMP-1, 8, 13, 18): key enzymes in matrix degradation. It contains Zn atom
in their active site and it is secreted by fibroblast to digest the plasma fibronectin, forming
fragment of fibronectin. Besides, it involved in embryonic development, reproduction,
tissue remodeling. It is capable of break down collagen type I, II and III and soluble protein.
- Gelatinase (MMP-2,9): matrix zinc-metalloproteinase that degrade denatured collagens
(gelatins) as well as other extracellular matrix proteins. It can be produced by fibroblast,
chondrocytes, endothelial cells, macrophages, neutrophils, T lymphocytes, …
- Stromelysins (MMP-3,10,11): activate other metalloproteinase (collagenase, gelatinase,
matrilysin); degrades collagen types II, II, IV, IX and X, proteoglycans, fibronectin, laminin,
and elastin
Role:
- Break down of both matrix and non-matrix proteins to remove them from the tissue to
serve the roles in morphogenesis, wound healing, tissue repair and remodeling in
response to injuries and in progression of diseases (atheroma, arthritis, cancer, and
chronic tissue ulcer)
Regulation:
- Gene expression: growth factors, hormones, inflammatory cytokines, UV irradiation,
cell-cell and cell-matrix contacts
- Endogenous inhibitor: a2-macroglogulin (in blood plasma/body fluids) and tissue
inhibitor of metalloproteinases (TIMPs) (in the tissues)
- Allosteric activators on prometaloproteinase activation