Important Concepts to Remember
Block 1 Unit 1
Introduction to Cell Biology
Maximum Resolving Power=0.22 µm
Human cells ~ 10 µm
Living cells can’t be seen under EM (black and white images)
Organelle
Smooth ER
Size
Rough ER
Golgi
Peroxisomes
0.2-1 µm
Mitochondria
Up to 7 µm
Lysosome
0.3-0.8 µm
Glycogen
10-40 nm
Function
Synthesis of steroids and
detoxification of drugs; Ca++
accumulation and release
Protein synthesis
Accepts proteins from RER for
modification
Catabolizes long fatty acid
chains
Produces ATP; contains
mitochondrial DNA and
enzymes
Contains acid hydrolases (has
an acidic lumen)
Storage form of glucose
Six abnormal functions in the cell:
1.
2.
3.
4.
5.
6.
Swollen mitochondria
Dilated ER
Lysosomal storage
Lipid accumulation
Cytoplasmic accumulation of glycogen
Hypoxic vacuoles
Introduction to Proteins Part 1
Buffers resist change in pH; a good example is the bicarbonate system in the blood
L isomer predominant in nature; counterclockwise NH3, COOH, R group, H
R group classification of aa
R group Classification
Nonpolar, aliphatic R groups (hydrophobic)
Aromatic R groups
Aliphatic hydroxyl R groups (polar aa)
Aliphatic sulfydryl (thiol) R group (polar aa)
Basic R groups
Acidic R groups (-charge at pH 7)
aa
Gly, Ala, Val, Leu, Iso, Met, Pro
Phe, Tyr, Trp
Ser, Thr
Cys
Lys, Arg, His
Asp, Glu, Asn, Gln
Plasma Membrane 1
Proteins define the unique functions of membranes, while lipids form the permeability barrier and
define the basic architecture.
Membrane Assymetry:
Exoplasmic is choline-containing
Cytoplasmic is amino containing
Introduction to Proteins 2
Disulfide bonds are the most common covalent modification (formation of disulfide bridges). Disulfide
bonds are formed by the oxidation of –SH side groups of a pair of cysteine residues to S-S linkage to
form cystine.
MW= # aa X molecular weight (in Da)
Average molecular weight of aa=110 Da(.11kD)
Cell Structure and Function
3 major structures found in cytoskeleton=actin filaments, intermediate filaments, microtubules
Actin filamants are 15% of the total protein in the cell and consist of alpha, beta, and lambda classes
ACTING BINDING PROTEINS
Acting-Bindin Protein
Fimbrin
α-actinin
Spectrin
Filamin
Dystrophin
Function
Binds actin filaments
Binds membrane with fibroblasts, found
in intestines
RBC membrane inner surface
Network structure; “leading edge” of
moving cell
Present
in
muscle
cell;
if
defective=muscular dystrophy
INTERMEDIATE FILAMENTS
Inermediate Filament
Acidic/Basic Keratins
Distribution
Epithelial cells
Desmin, GFAP, vimentin
Neurofilaments
NFM, NFH)
Lamins
Muscle,
glial
mesenchymal cells
(NF1, Neurons
Proposed Function
Tissue
strength
and
integrity
cells, Sarcomere organization,
integrity
Axon organization
Nucleus
Nuclear structure
organization
and
The outer membrane of the nucleus is continuous with the RER, while the inner membrane of
the nucleus is continuous with the nuclear lamins.
Actin: Globular monomers of G-actin; 1 ATP per monomer; polymer is 2-stranded helix; 6 nm
diameter
Tubulin: globular dimers of alpha and beta that line up end to end; 1 GTP per dimer; polymer is
hollow tube composed of 13 protofilaments
NPC=nuclear pore complexes, composed of nucleophorin proteins
Molecules larger than 60 kDa require other proteins which interact with nucleophorins for
transport
Plasma Membrane 2
Voltage gated ion channels: rely on selectivity filter and dehydration of ions
Ligand-gated ion channels: ligand binding results in conformational change of receptor (energetically
favorable)
“Passive transport”: membrane transport mechanisms that transport molecules down the EC gradient,
therefore not requiring energy
“Active transport”: depends on energy input for function
3 types of active transporters: P-type, ABC-type, F & V type
Blood
Blood=6-8% of body weight
Total blood volume: 5.5 L (6 quarts)
RBC Volume=45%
Average concentration of Hb: 13.6-17.2 g/dl (Men), 12.0-15.0 g/dl (Women)
Erythrocyte count: 4.3-5.9 X 106 mm3 (Men), 3.5-5.0 X 106 mm3 (Women)
Hematocrit: packed erythrocytes per unit volume in blood; 40-50% (Men); 35-45% (Women)
Total erythrocytes: 5,000,000 per mm3 blood
Total leukocytes: 7000 per mm3 of blood
Total platelets: 250,000 per mm3 of blood
LEUKOCYTES
Cell
Neutrophil
Lifespan
<1 week
%
60-70%
Eosinophil
<2 weeks
2-4%
Basophils
1-2 years
<1%
Lymphocytes
Few months to several
years
20-25%
Monocytes
Few days in blood,
several months in
connective tissue
3-8%
Function
Phagocytosis and
destruction of bacteria
Phagocytosis of
antigen-antibody
complex: destruction of
parasites
Similar to mast cells to
mediate inflammatory
response
T cells: cell-mediated
inflammatory response
B cells: humorally
mediated immune
response
Differentiate into
macrophage;
phagocytosis,
presentation of antigen
HUMORAL IMMUNITY
Ig
IgG
IgM
IgA
Function
Immunity against bacteria and
viruses in extracellular fluid
Immunity against bacteria and
viruses in extracellular fluid
Secreted by plasma cells in GI,
Percent
80%
5%
15%
respiratory tract, genitourinary
tract, breast milk
IgE
Defense against multicellular
parasites and allergic reactions
IgD
Fx not known
T cells need AG presentation by MHC (II for helper, I for cytotoxic)
<1%
<1%
MHC I found on all nucleated cells of the body
MHC II found only on macrophages, macrophage-like cells, and B cells
Epithelial Tissue
Type of Tissue
Simple squamous
Simple cuboidal
Simple columnar
Pseudostratified columnar
Stratified squamous keratinized
Stratified squamous non-keratinized
Stratified cuboidal
Transitional (distended and non-distended)
Gustatory
Olfactory
Stato-acustic
Germinal
Location
Endothelium of blood and lymphatic vessels,
mesothelium of peritoneal cavity
Ducts of glands, covering of ovary, kidney tubules
GI tract, gall bladder, parts of reproductive tract
uterus, efferent ductules, part of respiratory tract
bronchioles
Most of respiratory tract, trachea, epidymus, nasal
cavity
Epidermis
Wet inner surfaces, oral cavity, esophagus, vagina,
conjunctiva of eye
Ducts of sweat glands, pancreas, salivary glands
Lines most of urinary tract
Tongue
Nasal passageway
Covering inner ear
Lines seminiferous tubules of testis
The apical surface of epithelial tissue is capable of endocytosis, exocytosis, and transcytosis. Exocytosis
occurs at the apical surface for exocrine glands and at the basolateral surface in endocrine glands.
Distinguish between the terminal bar and the terminal web:
Terminal web: where actin filaments embed at the base of the microvilli
Terminal bar: place where two cells contact or attach to each other near the apical surface,
located on the lateral domain of a cell; runs in a zone all around the circumference of a cell
Note that cadherins are transmembrane proteins located in all three levels of the terminal bar.
Gap junctions (nexus) have a low electrical resistance, so they are responsible for the peristaltic
contraction of smooth muscle in the gut.
Note that the basal domain serves to connect the epithelium to the underlying CT. Hemidesmosomes
are in the basal domain, have attachment plaques on the cytoplasmic side, and contain integrin instead
of cadherins.
Desmosomes are located in the basolateral domain and contain cadherins, which are TM proteins that
help link cells to adjacent cells.
Type of Secretion
Mucus
Description
Thick and viscous
Serous
Mixed
Sebum
Ceruminous
Watery
Serous/mucus
Oily
Waxy
Mode of Secretion
Merocrine
Description
Most common; excretion by
exocytosis
Secretion is pinched off of cell
with part of cytoplasm and cell
membrane lost
Cell fills up with secretion, dies,
and becomes the secretory
product
Whole living cell is the secretion
Apocrine
Holocrine
Cytocrine
Gland
Goblet cell, sublingual salivary
gland
Pancreas, parotid, or sweat glands
Salivary glands
Sebaceous glands of skin
Ceruminous glands of external
auditory canal
Gland
Sweat glands, salivary glands,
pancreas, goblet cells
Lipid portion of milk of
mammary gland
Sebaceous gland of the skin
Ovary, testis
Introduction to Proteins 3
Beta-mercaptoethanol: reduces covalent disulfide bonds –S-S- (cystines) to sulfhydryls -SH + HS(cysteine); forms disulfide bridges
8M urea: disrupts noncovalent bonds; thought to weaken hydrophobic interactions and disrupt
H bonds
Protein folding is facilitated by: electrostatic interactions, disulfide bridges; thermodynamically
favored
Marginal stability of proteins is biologically advantageous
Affinity chromatography: separates and purifies proteins based on that protein’s specific
binding affinity for the molecule attached to the column matrix
best method of protein purification b/c you can increase the yield of your protein of
interest dramatically
Connective Tissue
Tissue
Mesenchymal
Mucous
Loose (areolar)
Dense irregular
Dense regular collagenous
Dense regular elastic
Reticular
Adipose
Blood
Cartilage
Location
Embryo and fetus
Umbilical cord
Loose packing around most organs and tissues,
surrounds blood vessels
Dermis, organ capsules, periosteum,
perichondrium
Tendons, ligaments, aponeurosis (tendonous
extension)
Ligamentum nuchae, flava
Lymphatic tissue, bone marrow
Omentum, subcutaneous fascia
Cardiovascular, hematopoietic tissue
Costal cartilage, trachea, pina, epiglottis
Lipoprotein lipase makes white adipocytes larger because it breaks down fatty acids, VLDL, and
chylomicrons to free fatty acids and glycerol, which are then taken up and stored by adipocytes as lipids;
stimulated by insulin
Insulin: acts on adipocytes to form triglycerides from glucose, increases uptake of glucose and
production of lipoprotein lipase
Hormone sensitive lipase makes white adipocytes smaller by breaking triglycerides down into free FA
that complex with albumin the blood for transport around the body; stimulated by leptin, epinephrine,
and norepinephrine
Leptin: hormone produced by adipocytes that targets the hypothalamus. Decreases food intake and
increases energy consumption.
Epinephrine/norepinephrine: stimulate hormone sensitive lipase to mobilize fatty acids
Thermogenin: mitochondrial membrane protein that permits the back flow of H+ instead of using them
for ATP production; this uncouples the oxidation process and leads to heat generation in brown
adipocytes
Cell
Type
Fibroblast
Fixed
Mesenchymal
Fixed
Reticular
Fixed
Adipocytes
fixed
Chondrocyte
Fixed
Osteocyte
Fixed
Blood cells
Fixed
Macrophages
Wandering
Mast cells
Wandering
Plasma Cells
Endothelial
Smooth Muscle Cells
Wandering
Associated
Associated
Pericytes
Associated
Function
Synthesis and maintain
extracellular matrix of CT
Synthesis and maintain
extracellular matrix of CT
Synthesis and maintain
extracellular matrix of CT
Synthesis and maintain
extracellular matrix of CT
Synthesis and maintain
extracellular matrix of CT
Synthesis and maintain
extracellular matrix of CT
Synthesis and maintain
extracellular matrix of CT
Phagocytic cells, antigen
presenting cells
Store mediators of the
immune response
Produce antibodies
Line the blood vessels
Surrounds the endothelium
and controls vessel size
Multipotential for vessel
growth and repair
As a monocyte leaves the blood vessel and travels to the extracellular matrix, it becomes a macrophage.
Macrophages derive from monocytes (from a cell line called granulocytic-mononuclear stem cell) but
monocytes/macrophages are in fact agranulocytic.
Granules of mast cells contain, among other things, histamine (too much causes anaphylaxis) and
leukotrienes (which cause release of cytokines)
Remember that collagen accounts for 20% of the protein in the body and 30% of the dry weight of the
body.
Remember: Hydroxylysine holds together the three α-polypeptide chains to form tropocollagen, and
fibrils are held together by hydroxyproline, which binds the tropocollagen together.
TYPES OF COLLAGEN
Type
I
Function
Resists tension; forms thick
fibers
II
Resists pressure; forms thin
fibers
Forms structural
framework of spleen, liver,
lymph nodes, smooth
muscle, adipose tissue
Does not form fibers; forms
meshwork of the lamina
densa of the basal lamina
to provide support and
filtration
III
Aka reticular fibers
IV
Location
Dermis, tendon, ligaments,
capsules of organs, bone,
dentin, cementum
Hyaline cartilage, elastic
cartilage
Lymphatic system, spleen,
liver, cardiovascular system,
lung, skin
Basal lamina
Collagen Formation:
Collagen is transcribed in the nucleus and translated in the ER, where Pro and Lys are hydroxylated,
procollagen is formed and then secreted via the Golgi.
Procollagen peptidase: cleaves procollagen to tropocollagen
Lysyl oxidase: links hydroxylysine molecules of adjacent tropocollagen molecules to form fibrils
Distinguish between Ehler’s Danlos Type VII (due to procollagen peptidase change) and Ehler’s-Danlos
Type IV (deficiency in Type III collagen)
Remember that elastic fibers= elastin + microfibrils. Elastin contains the unique enzymes desmosine and
isodesmosine.
Elastic fibers are formed like collagen fibers, but lysyl oxidase links tropoelastin (instead of
tropocollagen) to form elastin (instead of collagen).
Two other types of elastic fibers exist: oxytalan fibers and eulanin fibers.
Aggrecan is a free proteoglycan whose core protein is chondroiton sulfate and keratin sulfate.
An aggrecan aggregate is HA with hundreds of aggrecan molecules. It is found in cartilage and is much
more gel-like and compression resistant.
PROTEOGLYCANS
Proteoglycan
Type
Core Protein
Location
Aggrecan
Free
Perlacan
Syndecan
Free
Transmembrane
Fibroglycan
Transmembrane
Chondroiton sulfate and
keratin sulfate
Heparin sulfate
Heparin sulfate and
chondroiton sulfate
Heparin sulfate
Cartilage matrix
(compression resistant)
Basal lamina
One end in cytoplasm
and the other end in
the EMC
Binds where there is
collagen and fibronectin
(in basal lamina)
GLYCOPROTEINS
Type of Glycoprotein
Fibronectin
Location
Basal lamina
Laminin
Entactin
Tenascin
Chondronectin
Osteonectin
Basement membrane
Basement membrane
Basement membrane
Cartilage matrix
Bone matrix
Binds to:
Heparin sulfate and collagen
type IV
collagen IV and heparin sulfate
Laminin and collagen IV
Syndycan and fibronectin
Cell integrin to collagen II
Cell integrin to collagen I
Remember that basement membrane is a light microscope term, is located beneath the basal surface
epithelia, and has 3 layers at the EM level.
Basal lamina is an EM terms and there are two views as to what composes it. Dr. Crissman and our text
present that the basal lamina consists of the lamina lucida and the lamina densa.
LAYERS OF THE BASAL LAMINA
Layer
Lamina lucida
Lamina densa
Location
Next to cell membrane
Fuzzy band below lamina lucida
Lamina reticularis
Below lamina densa
Contains:
Integrins, laminins, and entactin
Collagen IV, perlacan (heparin
sulfate) and fibronectin
Reticular fibers (collagen III),
collagen IV and VII of anchoring
fibrils
Remember that the basement membrane acts as a macromolecular sieve.
Myoglobin and Hemoglobin
O2 is not very soluble in water, Mb helps with this
Mb increases O2 availability in muscles
Hb carries O2 from capillaries in lungs to capillaries in tissues
N=hill coefficient; a measure of the degree of cooperativity
N<1 negative cooperativity, N>1 positive cooperativity, N=1 no cooperativity
In the Monod model, the T and R forms are in equilibrium. T state and O2 release is favored when O2 is
low. R state and O2 binding is favored when O2 is high (O2 binding stabilizes the R state). O2 affinity
increases as fractional saturation increases.
In the Koshland model, the protein is in either the T or the R form with no ligand bound. The ligand
binding to one subunit can cause a conformational change in a neighboring subunit.
The general model combines features of Monod and Koshland. The Monod model describes Hb binding.
Homotropic effectors: bind to the same type of site as the ligand (ex: O2 binding to Hb)
Heterotropic effectors: bind to a site different from the ligand, but still induces a
conformational change
Examples: H+ binds to His to stabilize the T state and reduces the affinity for O2.
Salt bridges form when His 146 is protonated, stabilizes the T (low affinity) state. Known as the
Bohr effect.
2,3-BPG binds to the center of Hb, stabilizes the T state and reduces the affinity for O2.
Hb binds one molecule of BPG, which reduces the affinity for O2
BPG is negatively charged and is surrounded by positive charge when it binds to Hb
CO2 reacts with the N-termini to stabilize the T-state and reduces the affinity for O2.
N terminus can carry CO2 and carbamino Val forms a salt bridge with Arg to stabilize T state.
O2 binding promotes the release of CO2 from the N termini.
High concentrations of O2 in the lungs result in saturation of Hb with O2, ligation of the O2
binding site reduces the affinity for CO2 and promotes CO2 release (Haldane effect).
Cl binding reduces affinity for O2.
Chloride binds to the deoxy form, decreasing O2 affinity
Forms a salt bridge between Arg on one subunit and N-terminus of another subunit.
HB EFFECTORS
Ligand
Effect on O2 affinity
Effect of binding
O2
BPG
Increased
Reduces
CO2
Reduced
Protons
Reduced
Homotropic
Heterotropic effector, stabilizes
the T state
Heterotropic effector, stabilizes
the T state
Heterotropic effector, stabilizes
the T state
O2 affinity increases as temperature decreases. At lower temperatures, there is not a very high oxygen
requirement.
CO is toxic because it binds with a higher affinity to O2 site in Hb, so it physically blocks the O2 site and
causes higher O2 affinity so it can’t release O2 either. It also binds to cytochrome oxidase and myoglobin.
Cells in sickle cell disease aggregate when Val residues insert into hydrophobic pockets of another HbS
molecule, which leads to polymerization and aggregation and long fibers which cause RBC sickling.
TCA Cycle
“Waltz around the cycle”
-α-Ketoglutarateglutamate
Oxaloacetateaspartate
Citrateacetyl-CoAfatty acids
Succinyl CoAheme synthesis
Cell Motility
Two Phases: Extravasation and Diapedesis
Extravasation consists of 1)Activation of endothelial cells 2)Trapping 3)Adhesion 4)Migration through
wall
Diapedesid consists of 1)extension of lamellapodia 2)adhesion by focal attachments to adhesion
molecules and collagen fibers in ECM 3)cytoplasm flows forward 4)retraction with footprint
Laminin: regulates neural outgrowths
Fibronectin: promotes migration
Cell Cycle
Phase of cell cycle
G1
N
2n
Time
6-12 hours
S
G2
Mitosis
4n
4n
6-8
3-4
1 hr
Cancer cells have a different amount of genetic material than a diploid cell.
Cell cycle is regulated by:
1.
2.
3.
4.
5.
6.
7.
8.
Mechanical force (stretching)
Injury to tissue (ischemia)
Cell death
Growth factors or mitogens
Clock or timer (correct order of event)
Mechanism to ensure that each event is triggered only once per cycle
Completion of each step
Rate of progression
****Phosphorylation of p53 inhibits the cell cycle, while dephosphorylation of Rb inhibits the cell
cycle
p16 helps prevent cancer by cellular senescence=tumor suppression mechanism in which the levels
of p16 increase after a certain number of cell divisions to cause cell death; normal in aging but not a
good thing in cancer (i.e. in stem cells or beta cells in pancreas)
Loss of p53 allows cells with damaged DNA to survive and divide, thereby propagating potentially
oncogenic mutations
Enzymes
Enzymes have a higher affinity for the TS than for the substrate.
Catalysis works by: proximity, strain or distortion of a bond, acid-base catalysis, or nucleophilic
catalysis. Nucleophilic catalysts form a covalent complex with part of the substrate. Nucleophilic
groups (O of serine OH, S of Cys, N of Lys or His, COO-of Glu and Asp)
Chymotrypsin: hydrophobic residue of the substrate binds in the hydrophobic pocket on the
enzyme, His is in proximity with Ser and withdraws a proton (acting as a general base catalyst), Ser
now a good nucleophile, Asp H bonded to His and orients it with respect to Ser, Ser covalently binds
with the carbonyl carbon of the substrate, c terminal peptide fragment is released, water hydrolyzes
this
VITAMINS AS PRECURSORS
Vitamin
Cofactor
Reaction
Thiamin
TPP aldhehyde
Group transfer
Riboflavin
FMN or FAD
Electron/H transfer
Niacin
NAD or NADP
Electron/H transfer
Pantothenic
Coenzyme A
Acyl-group transfer
Pyridoxine
PyrodoxalP
Carboxyl transfer
Biotin
Biocytin
One-carbon reactions
Vit B12
Coenzyme B12
1,2 shifts of hydrogen
Lipoic acid
Lipolysine
H and acyl transfer
E-S formation is fast compared to E-S to product and enzyme, so it is the rate-determining step.
Michaelis-Menten Equation: 𝑉 =
𝑉𝑚𝑎𝑥•[𝑆]
𝐾𝑚+[𝑆]
Km=dissociation constant
Briggs-Haldane: Km is NOT a dissociation constant but it is a kinetic term.
Double reciprocal plot: can tell ratio of Km to Vmax, 1/Vmax (y intercept), Km (x intercept)
Competitive Inhibition: Km changes but Vmax does not
Noncompetitive Inhibition: Km stays the same but Vmax changes
Turnover number=Kcat; number of substrate molecules converted per molecule of enzyme per unit of
time