Foundation of Health and Disease
(FHAD 101 )
Solomon T. Gizaw (PhD)
Chapter 2
Amino acids, Peptides & Proteins
Contents
1.
AMINO ACIDS, PEPTIDES & PROTEINS
1. AMINO ACIDS
1. General structure
2. Classification of proteinogenic amino acids
3. Non-proteinogenic amino acids
4. Acid-base properties of amino acids
2. PEPTIDES AND PROTEINS
1. The peptide bond and its characteristics
2. Peptides of physiological significance
3. Functions and classifications of proteins
4. Structural organization of proteins
1. Primary structure
2. Secondary structure
1. alpha helix characteristics
2. stabilizing and destabilizing aa forces
3. parallel sheets and beta-turns
3. Tertiary structure
1. stabilizing forces
2. structure-function relationship of proteins
4. Quaternary structure
5. Protein denaturation
1. Definition
2. Denaturing agents
3/9/2024
Solomon T. Gizaw (PhD)
3. Effects of denaturation
(6hrs)
3
Amino Acids
 The chemical transformation of amino acids are distinct from those of
carbohydrates or lipids in that they involve the element nitrogen.
We must therefore examine the origin of nitrogen in biological systems and
its disposal.
Amino Acid Pool
… Dietary protein (100 g/day)
… Body protein (400 g/day)
… Synthesis of nonessential aa varies
Protein Turnover
… constant synthesis and degraded permitting
the removal of abnormal or unneeded proteins.
3/9/2024
Solomon T. Gizaw (PhD)
5
Amino acids
• Organic acids that contain one or more amino (–NH2) group(s).
• Structural units of proteins, obtained from them by hydrolysis.
• Perform important functions in their free form.
• A carboxylic group (–COOH), an amine group (–NH2) and a characteristic side chain,
radical or R-group (–R).
• Amphoteric electrolytes or ampholytes, i.e., react as proton donor acid by –COOH and as
proton accepting base by –NH2 to get negative or positive charges.
• Exceptions: Proline and hydroxyproline - have an unionizable imino group (–NH–).
• -amino acids are so-called because both of the primary –COOH and –NH2 groups are
attached to the same -carbon atom.
3/9/2024
Solomon T. Gizaw (PhD)
6
Functions of amino acids
• Polymerized to form proteins.
• Stabilize the 3-D structure of proteins by forming H and disulfide bonds.
• The presence of specific AAs at the active site of enzymes is vital for catalytic activity.
• Some AAs(glucogenic) can be converted to carbohydrates.
Cys and met are sources of S in the body.
C skeleton and N of aas used for the synthesis of purine and pyrimidine bases for
nucleotides and nucleic acids.
Gly and met help in the detoxification mechanisms.
Met can act as a methyl group donor in methylation reactions.
3/9/2024
Solomon T. Gizaw (PhD)
7
Certain AAs give rise to biologically important derivatives
• Gly is a precursor for ‘heme’ of hemoglobin
• Gly is also a precursor for creatine that acts as the mediator of energy in muscles.
• Tyr is the precursor for a number of hormones (Thyroxine, triiodothyronine,
epinephrine and nor-epinephrine) and skin pigment melanin.
• Tryptophan can give rise to vitamin niacin and reduce its dietary requirement.
• Tryptophan also gives rise to the neurotransmitter, Serotonin(C10H12N2O)
• His can be converted to the mediator of allergic reactions i.e. histamine.
3/9/2024
Solomon T. Gizaw (PhD)
8
Fate of amino acids in the fed state
Protein turnover
Glucose + Ketone
bodies
Oxidize
Amino Acids Hepatic portal vein
(intestine)
Liver
In liver,
Synthesis of





3/9/2024
Serum proteins
nonessential amino acids
heme, hormones
neurotransmitters, and
purine and pyrimidine bases (e.g.,
adenine and cytosine in DNA).
Solomon T. Gizaw (PhD)
9
Maintenance of the blood amino acid pool
3/9/2024
Solomon T. Gizaw (PhD)
10
Chemical Classification of AAs
 Nonpolar, Aliphatic Amino Acids
 Aromatic Amino Acids
 Aliphatic, Polar, Uncharged Amino Acids
 Sulfur-Containing Amino Acids
 The Acidic and Basic Amino Acids
3/9/2024
Solomon T. Gizaw (PhD)
11
Abbreviations for the Amino Acids Name
3/9/2024
Solomon T. Gizaw (PhD)
12
The side chains of the amino acids
3/9/2024
Solomon T. Gizaw (PhD)
13
Chemical Classification of AAs
• Based upon the number of amino groups and carboxyl groups in AA
• The three groups under this classification are:
a) Neutral AAs - mono-amino mono-carboxylic;
b) Acidic AAs - mono-amino dicarboxylic;
c) Basic AAs -
diamino (or complex amino) mono-carboxylic AAs.
• Neutral AAs -further classified into
Aliphatic AAs with a hydrophobic nonpolar hydrocarbon side chain. Gly, Ala, Val, Leu,
Ile, Pro, Met.
Hydroxy AAs have a –OH group on the side chain-Ser, thr, tyr.
Also include 4-hydroxyproline and 5-hydroxylysine - important for cross-binding of
polypeptides, particularly in collagen.
Selenocysteine present in a few proteins is derived from serine that is modified during
translation.
and
3/9/2024
Solomon T. Gizaw (PhD)
14
Chemical Classification of AAs
• Aromatic AAs - carry an aromatic ring:
Phe (hydrophobic), Tyr (hydrophilic, -OH) and Trp (weak hydrophobic, iodole ring).
• S-containing AAs -contain a S atom in the side chain and are sources of S in the body:
Cys (and cystine) , nonpolar methionine.
• Heterocyclic AAs- contain heterocyclic ring structures:
His with a basic imidazole ring;
Trp with an iodole ring; and
Pro and Hydroxyproline with 5-membered intramolecular ring
3/9/2024
Solomon T. Gizaw (PhD)
15
Acidic & basic AAs
• Acidic AAs: Glu, Asp.
• Also occur in the body as amides by amidation of their side chains e.g., Glu is amidated
into Gln and asp into Asn.
Gln - a major role in removing NH3 from the brain.
Glu per se and as γ-amino butyric acid - from which it is synthesized - is a
neurotransmitter and is essential for urea and glutathione synthesis.
• Asn - protein cross-linking and covalent modification by glycosylation.
3/9/2024
Solomon T. Gizaw (PhD)
16
Basic AAs
• Include lysine, histidine and arginine, and the non-protein AAs, ornithine and citrulline.
• Ornithine and citrulline: Have no specifying genetic codons but they are metabolically
essential as free AAs. They both give rise to the basic guanido group.
• Nitric oxide, a hormonal second messenger is synthesized from Arg.
• Ornithine -used for the synthesis of polyamines that are important for cell cycle control.
• Derivatives of Lys found in proteins include; 5-OH lysine, 6-NH-CH3 lysine and the
complex desmosine derived from four lysines found in the fibrous protein elastin.
3/9/2024
Solomon T. Gizaw (PhD)
17
Biological classification of AAs
3/9/2024
Solomon T. Gizaw (PhD)
18
Biological classification of AAs
• Based upon whether the AAs can be synthesized in human body or not .
Indispensable or essential AAs - not synthesizable in the body in adequate amounts
and must be supplied in the diet.
e.g. Val, Ile, Thr, Trp, Arg, Leu, Lys, Met, Phe, His
Dispensable or non-essential AAs - synthesizable in the body and there is no diet
dependency for them.
e.g. Glu, Gln, Asp, Asn, Gly, Ala, Pro, Tyr, Ser , Cys.
• Biologically and nutritionally, the dietary deficiency or imbalance of any of the essential
AAs leads to nutrition deficiency disorders that affect both growth and health.
• Arg and His are semi-essential : The healthy adult human body synthesizes just enough
Arg and His but such amount is not enough in the childhood growth period, sickness,
convalescence and during pregnancy and requires dietary supplementation.
3/9/2024
Solomon T. Gizaw (PhD)
19
• Some AAs become essential under certain conditions:
Tyr also turns essential upon dietary deficiency of Phe;
Cys turns essential upon dietary deficiency of Met.
• The main dietary source of essential AAs is proteins of high biological value i.e.,
digestible proteins that contain all of them in well-balanced proportions.
These proteins include all animal proteins (e.g., milk, egg, meat, liver, fish and
chicken) and a few plant proteins (e.g. from beans and lentils).
• The remaining non-essential AAs are diet independent because they are synthesized in
the body, but they are metabolically as important as the essential ones.
3/9/2024
Solomon T. Gizaw (PhD)
20
Metabolic classification of AAs
3/9/2024
Solomon T. Gizaw (PhD)
21
Metabolic classification of AAs
Based upon the catabolic fate of C skeleton of AAs.
• Glucogenic AAs: 14 of the 20 protein AAs give
rise to intermediates of glycolysis or Kreb’s
cycle and thus can be converted to
carbohydrates, hence called glucogenic.
• Ketogenic AAs: Only two AAs are purely
ketogenic viz. lysine and leucine -catabolically
give intermediates convertible into acetyl-CoA
or acetoacetyl-CoA.
• Mixed AAs: These are AAs, the C skeleton of
which is catabolized to produce the glycolytic
intermediates as well as the acetyl -CoA
derivatives.
3/9/2024
Solomon T. Gizaw (PhD)
22
Catabolism of the Carbon Skeleton of Amino Acids
• The pathways by which amino acids are catabolized are conveniently organized according
to which one (or more) of the seven intermediates listed above is produced from a
particular amino acid.
1. Amino acids that form oxaloacetate
2. Amino acids that form α-ketoglutarate
3. Amino acids that form pyruvate
4. Amino acids that form fumarate
5. Amino acids that form succinyl CoA
6. Amino acids that form acetyl CoA or acetoacetyl CoA
3/9/2024
Solomon T. Gizaw (PhD)
23
Unusual/ modified AAs: Selenocysteine
• Some AAs after their incorporation into the proteins, are modified by hydroxylation,
methylation or carboxylation to form unusual AAs,
e.g. hydroxyl-proline, hydroxyl-lysine, γ-carboxy glutamic acid.
• Selenocysteine(Sec): Se is substituted in place of S of Cys to form selenocysteine.
--- This AA (called as 21st AA of proteins) is present at the active site of several
enzymes: e.g. gluthathione peroxidase,
Se has long been associated with antioxidant activity.
A glycoprotein has been isolated from mammalian blood that contains as many as 10
selenium residues.
Selenocysteine (Sec) is formed during translation and a special tRNA specific for
‘UGA’ (STOP) codon is used.
3/9/2024
Solomon T. Gizaw (PhD)
24
Unusual/ modified AAs: Pyrrolysine
• Pyrrolysine: Recently (2002), Pyrrolysine, has been detected which is encoded by another
stop codon, ‘UAG’, in several methanogenic archaea and bacteria organisms.
• Pyrrolysine, hence, is termed as the 22nd protein AA present in several methyltransferase enzymes.
• Pyrrolysine is attached to tRNA CUA by pyrrolysine-tRNA synthetase (a class II
aminoacyl-tRNA synthetase).
• The enzyme uses ATP to activate Pyrrolysine and ligates it to tRNA carrying CUA as the
anticodon that results in the translation of UAG as a sense codon rather than STOP.
3/9/2024
Solomon T. Gizaw (PhD)
25
Proteinogenic vs Non-Proteinogenic Amino acids
3/9/2024
Solomon T. Gizaw (PhD)
26
Proteinogenic vs
• Proteinogenic(genetically encoded) amino acids are amino acids that are incorporated
biosynthetically into proteins during translation.
• The word "proteinogenic" means "protein creating".
• 22 AAs are polymerizable into protein structure - protein AAs.
 20 in the standard genetic code and
 an additional 2 (selenocysteine and pyrrolysine) that can be incorporated by
special translation mechanisms
• Those that do not occur in proteins - non-protein AAs.
3/9/2024
Solomon T. Gizaw (PhD)
27
Non-Proteinogenic Amino acids
• Non-proteinogenic amino acids are amino acids that are either
not incorporated into proteins (like GABA, L-DOPA, or triiodothyronine),
misincorporated in place of a genetically encoded amino acid, or
not produced directly and in isolation by standard cellular machinery
(like hydroxyproline).
o The latter often results from post-translational modification of proteins.
• Some non-proteinogenic amino acids are incorporated into nonribosomal peptides which
are synthesized by non-ribosomal peptide synthetases.
3/9/2024
Solomon T. Gizaw (PhD)
28
Non-Protein AAs
• Some D-AAs have been found to play some role in biological systems.
o D-Ala and D-Glu are present in certain bacterial cell walls.
o D-Asp and D-Ser have been isolated from brain tissue.
• The AAs that do not exist in the proteins, perform very important functions as :
- - Biologically active molecules like hormones or neurotransmitters,
--- Important structural components of biomolecules like Coenzyme A, Acyl carrier
proteins etc. or are
- - intermediates in the metabolic pathways.
3/9/2024
Solomon T. Gizaw (PhD)
29
Acid-base properties of amino acids
3/9/2024
Solomon T. Gizaw (PhD)
30
Properties of AAs
• Stereoisomerism: -AAs (except glycine) have at least one asymmetric (chiral) C - show
stereoisomerism due to differential orientation of the amino and carboxylic groups in
relation to the side chain.
• All AAs (except glycine) are optically active, i.e., can rotate plane polarized light.
• Serine is used as a reference for configuration of AAs as one of the enantiomers;
• L-form with –NH2 on the left side of -carbon, is the predominant form, the only
form of AAs that occurs in proteins and is metabolizable ; and the D-form, that is
unmetabolizable with –NH2 on the right side of -carbon.
3/9/2024
Solomon T. Gizaw (PhD)
31
• D-AAs are found in the peptides of bacterial cell walls and certain antibiotics.
• Aromatic AAs (Trp, Tyr and Phe) can absorb UV light at 280 nm; a property utilizable for
identification and quantization of AAs & proteins in solution.
3/9/2024
Solomon T. Gizaw (PhD)
32
Stereoisomerism of AAs : a) Ball and stick model b) Howarth projections
3/9/2024
Solomon T. Gizaw (PhD)
33
Ionization and Isoelectric point
• Carboxylic as well as the amino groups of AAs are ionizable.
• In alkaline pH, the carboxylic ‘–COOH’ group of an AA (free or protein-bound) gets
dissociated to release a proton and acquires a –ve charge, whereas, the –NH2 group ionizes
in acidic pH into + ve charged amino –NH3+.
R-COOH  R-COO- + H+
R-NH3+  R-NH2 + H+
• The ionization of all the groups is pH specific i.e. each functional group is protonated or
deprotonated at a particular pH called pKa.
• At extremely alkaline pH, the AAs would carry a net negative charge whereas, in acidic
conditions, they will be positively charged.
3/9/2024
Solomon T. Gizaw (PhD)
34
Isoelectric pH
3/9/2024
Solomon T. Gizaw (PhD)
35
• Isoelectric pH:
The specific pH at which an AA (or a protein) would carry equal negatively charged –COOand positively charged –NH3+ groups is called isoelectric pH or isoelectric point (pI).
• In this state the molecule carries no net charge or is ‘isoionic’ and is called Zwitterions
(meaning ‘equal charge ion’).
• Since at pI, the AAs are electrically neutral, they will not migrate in electric field and are
readily precipitated.
• The pI is a specific characteristic for each AA or protein, e.g., for alanine, it is 6.02 and for
aspartic acid, it is 2.98 .
• This characteristic is utilized for protein separation by electrophoresis and identification
by isoelectro-focusing.
3/9/2024
Solomon T. Gizaw (PhD)
36
Titration of glycine: The ionic species of glycine at different dissociation points
3/9/2024
Solomon T. Gizaw (PhD)
37
Peptides & Proteins
• Peptides: Short chain of amino acids linked by peptide bond.
• Polypeptide: a longer, continuous, unbranched peptide chain.
• Protein: Polypeptides which have a molecular mass of 10,000 Da or more.
3/9/2024
Solomon T. Gizaw (PhD)
38
Peptides of biological importance
• Bradykinin :
 local anti-inflammatory, vasodilator, hypotensive hormone, potent smooth muscle
relaxant , formed of 9 AAs. (NH2-Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg-COOH).
• Oxytocin (love drug/hormone)
- a posterior pituitary hormone that induces uterine contraction, nonapeptide.
CYIQNCPLG-NH2
• Thyrotropin-releasing hormone (TRH) :
- Tripeptide composed of a cyclized Glu (pyro-glutamic acid), His, and prolinamide;
the hypothalamic hormone responsible for the stimulation of the pituitary to secrete
thyrotropin (TSH)
the carboxamide derivative of L-proline
3/9/2024
Solomon T. Gizaw (PhD)
39
Physiological Systems involved in a thyroid hormone homeostasis
–
–
–
–
–
–
–
–
–
–
D1/2/3 deiodinase 1/2/3,
T2 diiodothyronine,
rT3 reverse triiodothyronine (inactive),
RXR retinoic acid receptor,
TRE thyroid hormone response element,
T3 triiodothyronine,
T4 thyroxine,
THR thyroid hormone receptor,
TRH thyrotropin-releasing hormone,
TSH thyrotropin (thyroid-stimulating
hormone),
– PVN periventricular nucleus (of the
hypothalamus)
3/9/2024
Solomon T. Gizaw (PhD)
40
• The major circulating TH is thyroxine (T4), whose synthesis is limited to the thyroid gland.
• T4 is considered a pro-hormone, which, at the local level, is regulated by tissue-specific
ways by transformation into activated (triiodothyronine [T3]) or inactivated (reverse-T3
[rT3]) metabolites.
• Three different deiodinases have been described: iodothyronine deiodinase 1 (DIO1),
DIO2, and DIO3.
3/9/2024
Solomon T. Gizaw (PhD)
41
Glutathione
• Tripeptide (-glutamyl-cysteinyl-glycine), coenzyme, antioxidant.
• It participates in a number of important metabolic reactions e.g. detoxication, eicosanoid
synthesis and transport of AAs across cell membranes.
3/9/2024
Solomon T. Gizaw (PhD)
42
Pathogenesis
• G6PD catalyzes NADP reduction into NADPH.
• NADPH protects cells from oxidative damage by regenerating reduced glutathione (GSH ).
• Erythrocytes do not generate NADPH in any other way, they are more susceptible than other cells
to destruction from oxidative stress.
3/9/2024
Solomon T. Gizaw (PhD)
43
Role of the PPP in the Generation of NADPH
• The glutathione-mediated defense against oxidative stress is common to all cell types (including
the RBC), and the requirement for NADPH to maintain levels of reduced glutathione probably
accounts for the universal distribution of the pentose phosphate pathway among different types
of cells.
3/9/2024
Solomon T. Gizaw (PhD)
44
• Insulin : formed from 2 polypeptide chains connected by 2 disulfide
linkages; one is 30 and the other is 21 AA residues.
3/9/2024
Solomon T. Gizaw (PhD)
45
Insulin is a polypeptide hormone of 51 amino acids that is composed of two
polypeptide chains connected by 2 disulfide linkages; one is 30 and the other is 21
AA residues
3/9/2024
Solomon T. Gizaw (PhD)
46
• Beta-lipoprotein : hypophyseal hormone that stimulates the release of FAs from adipose
tissue, large polypeptide (91 AAs)
a lipoprotein that transports cholesterol in the blood;
composed of moderate amount of protein and a large amount of cholesterol;
high levels are thought to be associated with increased risk of coronary heart disease
and atherosclerosis
3/9/2024
Solomon T. Gizaw (PhD)
47
Proteins: structure and properties
3/9/2024
Solomon T. Gizaw (PhD)
48
Proteins: structure and properties
• High MW, formed of C, O, H & N.
• May also contain S, P, non-protein organic groups and metal ions.
• Polymers of subunits of the polymerizable, genetically coded 20 (now 22) AAs linked
together by peptide linkages.
• May be monomeric or polymeric.
• Proteins: simple, complex and derived
Simple proteins formed of AAs only .
Some proteins have a non-proteinaceous moiety attached covalently or noncovalently to the polypeptide chain(s) → Complex or conjugated proteins.
3/9/2024
Solomon T. Gizaw (PhD)
49
49
Proteins: structure and properties
• Alteration in AA composition/sequence of proteins →
change in the
conformation or stability of the proteins, thus compromising their cellular
function. Such alterations originating at the genetic level lead to clinical disorders
known as inherited disorders.
• In the human body, there are ~100,000 different proteins .
3/9/2024
Solomon T. Gizaw (PhD)
50
Bonds participating in the protein structure
 Peptide bond
 Disulfide bond
 H-bond
 Electrostatic interaction
 Van der Waals interactions
 Hydrophobic interactions
3/9/2024
Solomon T. Gizaw (PhD)
51
Bonds participating in the protein structure
• Peptide bond: AAs are polymerized to give rise to polypeptides or proteins by the
formation of peptide bonds. It imparts restriction in the rotation around C-N bond
leading to specific type of folding patterns called secondary structures.
•
The peptide bond is the strongest bond in protein structure.
• Disulfide bond: The side chain –SH of two cysteine residues of a protein join with the
removal of two H atoms (oxidation) to form a disulfide bond. It may be intra-chain or
inter-chain. Weaker than peptide bond in strength because it can not resist denaturation.
3/9/2024
Solomon T. Gizaw (PhD)
52
Bonds participating in protein structure
• Hydrogen bond: Weak partial electrostatic attraction between
polar molecules through the participation of H atom.
• Although weak, the presence of H-bonds in large numbers plays
a critical role in stabilizing specific secondary structures in
proteins as well as nucleic acids.
• H-bond is formed when H-atom is shared between two highly
electronegative atoms (e.g. N, O, or P).
• Polarization of H bonds confers polarization to the molecules
that become consequently hydrophilic, i.e., soluble in H2O.
• Electrostatic interactions: Large differences in electronegativity of
participating atoms generate weaker non-covalent electrostatic/
ionic /salt bonds due to unequal sharing of electrons.
3/9/2024
Solomon T. Gizaw (PhD)
53
Bonds participating in protein structure
• Van der Waals interactions: The weakest type of bonds.
• Within each molecule, atoms are surrounded by charges that vary asymmetrically
and temporally in distribution.
• Asymmetricity within the molecule induces similar changes in neighboring molecules
that create dipolar interactions between them known as the van der Waal forces.
• Such nonspecific forces could be attractive when molecules are an optimal van der
Waal distance apart or could be repulsive when molecules are too close to each
other.
• Although weak, they play an important role in intra- and inter-molecular protein
stability.
3/9/2024
Solomon T. Gizaw (PhD)
54
Bonds participating in protein structure
• Hydrophobic interactions: The nonpolar groups, e.g. hydrocarbon side chains of AAs, in
an aqueous environment tend to cluster tightly to minimize the hydrophobic surface so
as to avoid interaction with polar water molecules.
• The water-hating property of non-polar molecules rather than their affinity to one
another holds them together.
3/9/2024
Solomon T. Gizaw (PhD)
55
Structural organization of Proteins
3/9/2024
Solomon T. Gizaw (PhD)
56
3/9/2024
Solomon T. Gizaw (PhD)
57
Structural organization of Proteins
• Polypeptides generally have MW below 10,000, whereas, proteins have higher MW.
• The average MW of an AA residue in a protein is ~ 110 Da and therefore, the average
MW of a protein = 110 x N, where ‘N’ is the number of AA residues.
• The Function of monomeric or polymeric proteins is determined largely by their
sequence, structural conformation, and nature of interaction among its structural units:
sequence-structure-function relationship.
3/9/2024
Solomon T. Gizaw (PhD)
58
Levels of organization of protein structure
• Primary structure: Specifies the total number of
AAs; the sequential linear order of composite AAs
connected by peptide bonds, and relative
participation of each AA in the peptide chain.
• Also specifies the possible position of intra- and
inter-molecular disulfide bonds, if present.
• Conventionally, peptide sequence is written left to
right.
• The left side terminal AA is the peptide’s Nterminal and the final AA towards the right end
is the peptide’s C-terminal.
3/9/2024
Solomon T. Gizaw (PhD)
59
Secondary structure
• Specifies the fine conformation of peptide
backbone created by spatial arrangements
and interactions of AA residues that are near
one another in the linear sequence.
• α-Helix: The most common secondary
structure, about ¼ of all AA residues in
polypeptides are found in α-helices.
3/9/2024
Solomon T. Gizaw (PhD)
60
β-pleated sheets& β bends
 β-pleated sheets: Polypeptides (2-5 chains) run in
line with each other and H bonding stabilizes the
structure.
 When the polypeptide chains run in parallel direction
to each other, called ‘parallel β-pleated sheets’ and
when they run in opposite directions to each other,
called ‘anti-parallel β-pleated sheets’.
3/9/2024
Solomon T. Gizaw (PhD)
61
Triple helix & Supersecondary structure
• Triple helix: Seen in collagen fibres which are rich in pro and hydroxyproline along with
lys. The 3 polypeptide strands are wound around each other.
• The triple helices running parallel are cross linked to form very stable structures.
• The triple helix is stabilized by inter chain H bonds as well as noncovalent interactions.
3/9/2024
Solomon T. Gizaw (PhD)
62
Triple helix & Supersecondary structure
• Super secondary structures: A number of proteins show the formation of super
secondary structures by a combination of more than one type of secondary structures.
a) β-α-β units: Two β units separated by an α-helix .
b) The Greek key: Several β sheets join to form a structure often found on classical
Greek pottery items.
c) β-meander: 5 polypeptides with β-pleated sheet structures joined by β-turns run
antiparallel to each other to form β-meander.
The β-sheets are stabilized by extensive H bonding.
3/9/2024
Solomon T. Gizaw (PhD)
63
Tertiary structure
 The
spatial
arrangement
and
interrelationship of various regions/ domains
is known as its tertiary structure.
 Specifies the final functional 3D structure of a
protein formed by complicated folding and
super-folding of the peptide chain into
globular or fibrous form of different size.
 Stabilized by interactions through R-groups of
composite AAs utilizing van der Waals, ionic,
hydrophobic, H and disulfide bonds.
 As it is the biologically active conformation of
the polypeptide, is the most labile to
denaturation.
3/9/2024
Solomon T. Gizaw (PhD)
64
Quaternary structure
• Specifies the nature of the association of more than one polypeptide in a defined
geometric positional configuration to make a functional protein.
• The polypeptides in a quaternary protein could act in synergy, suppress or stimulate the
function(s) of each other.
• Many proteins contain 2, 3, 4 or more subunits: dimeric, trimeric, tetrameric or
oligomeric proteins.
• Polymeric proteins could be hetero-oligomers (i.e., several polypeptides each with
different sequence) or homo-oligomers (i.e., several polypeptides of the same type).
• Oligomeric proteins are very common among enzymes e.g. carbonic anhydrase is
composed of 27 polypeptides.
• Quaternary structure stabilized by all types of secondary non-covalant bonds and is
the most sensitive to denaturation.
3/9/2024
Solomon T. Gizaw (PhD)
65
Quaternary structure
 Many times. the protein structure alternates between two functional states that may get
interchanged upon binding to other proteins or allosteric low MW effectors. Such
proteins are allosteric proteins, e.g., Hb (tetrameric) and changes its conformation upon
binding with O2 or CO.
 Cooperativity : one of the major characteristics of quaternary structure. The different
subunits of oligomeric proteins exchange information and affect the functional
characteristics of each other. The protein has +ve cooperativity if the activity is enhanced
by the binding of a modulator and –ve cooperativity if the activity is decreased.
3/9/2024
Solomon T. Gizaw (PhD)
66
Quaternary structure
• The proteins, at their final tertiary or quaternary conformation, may be aggregated into
functional macro-molecular complexes
e.g.
cell membrane proteins,
electron transport proteins and
metabolic enzyme complexes, e.g., fatty acid synthase enzyme.
• In such superstructures, abnormality in one or more of the composite proteins affects
the whole structure-function efficiency.
3/9/2024
Solomon T. Gizaw (PhD)
67
Classification of Proteins
• About 26,000 to 40,000 genes in human genome.
• Different criteria for classification:
biological value,
axial ratio
function
physical properties.
• Nutritional value: Nutritional value of proteins depends upon their digestibility and AA
composition.
 High biological value when they are digestible and contain all the essential AAs in
well-balanced proportions; or
low biological value when they are deficient in one or more AAs or contain them in a
very low imbalanced amount or are indigestible.
3/9/2024
Solomon T. Gizaw (PhD)
68
68
Classification of Proteins
• Proteins of high biological value :
all digestible proteins of animal origin ( dairy products like eggs, liver, fishes, red and
while meats) and
some proteins of plant origin ( lentils and broad beans).
• Most plant proteins and a few animal proteins are of low biological value
 e.g. collagen deficient in trp and cys ;
skin, hair, and nail keratins are indigestible.
• However, a protein of high biological value is not a dietary must because different low
biological value sources can complement each other and compensate for deficient AAs.
3/9/2024
Solomon T. Gizaw (PhD)
69
Classification of Proteins
• Axial Ratio: Proteins could be divided into globular or fibrous, depending upon their axial
ratio i.e. their molecular length: width indicated by ultramicroscopic studies.
• Fibrous proteins: Proteins with an axial ratio of >10 are fairly resistant and strong →
fibrous proteins.
• Majority have structural function e.g. keratins in hairs, wool, skin, nails and
cytoskeletons and myosin of the muscles.
• Globular proteins: The proteins with an axial ratio <10
• Less stable than fibrous proteins and more labile to denaturation.
• Most globular proteins are metabolically active and function as enzymes,
hormones or transport proteins e.g. Chymotrypsin, Igs, albumin, Hb and insulin.
3/9/2024
Solomon T. Gizaw (PhD)
70
Functions of proteins
• Structural- Main structural component in bone, muscles, cytoskeleton and cell
membrane.
• Nutrition- Provide the body with essential AAs, N and S.
• Catalytic- All metabolic enzymes are proteins in nature.
• Endocrine- Most hormones and all receptors are protein in nature.
• Defence: The antibodies (immunoglobulins) and complement systems that play an
important role in the body’s defensive mechanisms are proteins in nature.
• Osmotic Potential- Plasma proteins are responsible for the most effective osmotic
pressure of the blood. This osmotic pressure plays a central role in many processes, e.g.,
urine formation.
• Blood clotting factors are proteins.
3/9/2024
Solomon T. Gizaw (PhD)
71
Functions of proteins
 Transport - Proteins carry lipids in the blood forming lipoprotein complexes. Proteins
also carry, hormones, e.g., thyroid hormones and minerals, e.g., Ca, Cu & Fe.
Hemoglobin (a chromo-protein) carries O2 from the lung to tissues.
 Membrane transport:- The plasma membrane is lipoprotein and is semipermeable in
nature. The proteins in the membranes act as channels or transporters to allow
selective molecules/ions to cross into or out of the cells.
 Gene Regulation- Control of cellular activities through control of gene expression: Most
factors required for DNA replication, transcription and mRNA translation are protein in
nature.
 Signal Transductionlargely by proteins.
3/9/2024
Intercellular and intracellular signal transduction carried out
Solomon T. Gizaw (PhD)
72
72
Structure, Solubility and physical properties
• Simple proteins: The proteins composed of AAs only - Insulin, Myosin, Albumin etc.
• Conjugated proteins: The proteins that also carry certain non-AA prosthetic moieties like
heme, carbohydrates, lipids, nucleic acids, etc.
• Conjugation with prosthetic groups is stabilized by covalent and non-covalent
secondary bonds and one protein could contain more than one type of such groups.
• Derived Proteins: All the proteins derived from either of the above two types.
• Partial hydrolysis products of proteins (e.g., proteoses, peptones, long and short
peptides) and denatured proteins are derived proteins.
3/9/2024
Solomon T. Gizaw (PhD)
73
Protein Denaturation
3/9/2024
Solomon T. Gizaw (PhD)
74
Protein Denaturation
• Loss of native form with disruption of sec., tert. & quat. structure
changes in physical and chemical characteristics
 loss of biological activity
• No hydrolysis of peptide bonds, primary structure preserved.
• Factors disrupting weak secondary bonds responsible for maintaining protein structure:
physical manipulations: shaking, high temperature, and ionizing irradiations;
chemical factors e.g. organic solvents like acetone, strong alkalis, and acids; agents
that irreversibly precipitate proteins such as alkaloidal reagents and heavy metals,
and formaldehyde, formamide, glyoxal, SDS, guanidine hydrochloride and
isothiocyanate.
Some biological factors e.g. insulinase disrupts interchain disulfide bond of insulin
and inactivates it.
3/9/2024
Solomon T. Gizaw (PhD)
75
Properties of Proteins – Denaturation
• Changes in physical properties upon denaturation :
↑ viscosity,
↓ solubility
↓ diffusibility
• Denatured collagen: exception as denatured collagen (i.e. gelatin) is more water soluble.
• Chemical changes: loss of H, hydrophobic and electrostatic bonds, and exposure of
otherwise hidden groups.
This makes denatured proteins antigenically and immunologically distinct from their
native form.
• Impairment of structure-function relationship → Loss of enzymatic, hormonal, and other
biological activities.
3/9/2024
Solomon T. Gizaw (PhD)
76
76
Advantages of denaturation
• There are some advantages of denaturation  Proteins denatured by cooking are more easily digested than native proteins;
 Heat coagulation (denaturation) test is used for detection of urinary albumin;
 Measurement of specific analytes in biological samples such as uric acid and glucose
requires removal of proteins that could interfere with the estimation.
• So the blood/serum sample is first treated with alkaloidal reagents to denature
and precipitate the proteins.
Several traditional medical approaches for stoppage of bleeding and treatment of
burns are based on precipitation and denaturation of a superficial protein layer
such as alum, coffee powder and picric acid.
Denaturing conditions in electrophoresis are used to separate the proteins based on
MW
3/9/2024
Solomon T. Gizaw (PhD)
77
Prion Disease
3/9/2024
Solomon T. Gizaw (PhD)
78
Prion Disease
 Prion: "proteinaceous infectious particle”
 A prion disease is a type of proteopathy or disease of structurally abnormal proteins.
 Prion diseases are rare and occur due to proteins in the brain that “misfold.”
 aka transmissible spongiform encephalopathies (TSEs), fatal neurodegenerative disease in
humans and animals.
3/9/2024
Solomon T. Gizaw (PhD)
79
Prion Disease
• Prion disease represents a group of conditions that affect the nervous system in humans
and animals.
• In people, these conditions impair brain function, causing changes in memory,
personality, and behavior; a decline in intellectual function (dementia); and abnormal
movements, particularly difficulty with coordinating movements (ataxia).
• The signs and symptoms of prion disease typically begin in adulthood and worsen with
time, leading to death within a few months to several years.
• “Prions” are the disease-causing agents that can stimulate the abnormal folding of “prion
proteins.” In their usual, healthy state, prion proteins are typically present in the brain.
3/9/2024
Solomon T. Gizaw (PhD)
80
Prion Disease
• When prion proteins begin to fold abnormally and clump together — called amyloid
plaques — it leads to brain damage.
• They are all characterized by the accumulation of a protease-resistant isomer (PrPSc) of PrPC
in the brain of affected individuals.
• The normal form of the protein is called PrPC, while the infectious form is called PrPSc –
the C refers to 'cellular' PrP, while the Sc refers to 'scrapie', the prototypic prion disease,
occurring in sheep.
o Scrapie is a fatal, degenerative disease affecting the nervous systems of sheep and
goats.
3/9/2024
Solomon T. Gizaw (PhD)
81
Transmissible spongiform encephalopathies (TSEs)
• Creutzfeldt-Jakob disease (CJD): This type of CJD is split into three types: familial, sporadic,
and acquired.
• People inherit the familial type, but the sporadic type develops without any known
causes.
• Sporadic CJD is the most common type of CJD and tends to affect those aged around
60 years.
• A person can develop acquired CJD after unsterilized medical equipment has
introduced prions into the body, though this is rare.
• Variant Creutzfeldt-Jakob disease (vCJD): This type of CJD is an infectious prion disease
related to mad cow disease.
• People acquire it by eating meat containing proteins from the brain or spinal tissue of
a sick cow.
• Unlike sporadic CJD, vCJD is more likely to affect younger people.
3/9/2024
Solomon T. Gizaw (PhD)
82
Transmissible spongiform encephalopathies (TSEs)
• Fatal familial insomnia: This type — typically hereditary — is linked to inheriting an atypical
form of a gene that codes for prion proteins.
• Rarely, this disease occurs sporadically.
• Over the course of the disease, people sleep less and less.
• This can lead to mental deterioration and physical symptoms.
• Gerstmann-Straussler-Scheinker syndrome: This is a genetic disease that affects prion
proteins in the cerebellum.
• The cerebellum is the part of the brain that controls movement and balance, among
other functions.
3/9/2024
Solomon T. Gizaw (PhD)
83
The End of Chapter 2
3/9/2024
Solomon T. Gizaw (PhD)
84