Proteins are polymers of amino acids that are involved in most body functions and life processes. They have a primary structure defined by their amino acid sequence and levels of organization including secondary structures like alpha helices and beta sheets, tertiary structure describing the overall 3D shape, and potentially quaternary structure for multi-chain proteins. Proteins can be classified based on their function, solubility, composition and structure. Protein folding and structures are stabilized by various bonds and interactions.

1. Proteins
Dr. M.L. Mini
Associate Professor
Agricultural College & Research Institute
Madurai

2. Introduction
 Proteins are polymers of amino acids
 They are found in every cell in the body
 They are involved in most of the body’s functions and life processes
 proteins are linear chains of amino acids joined by peptide bonds
 Each protein has a characteristic amino acid sequence
Ala-Gly-Tyr-Trp-Arg-Cys-Gly-Pro-Ala-Glu-Asp-Gly
 Peptides are short polymers of amino acids
 Polypeptides contain many amino acids
 Protein may have one or more polypeptide chains

3. Classification
Based on function
 Catalytic proteins – Enzymes
 Regulatory proteins - Hormones
 Defence proteins – Immuoglobulins (antibodies)
 Storage proteins – globulins, prolamines and
glutelins of cereals, albumin in egg
 Transport protein – hemoglobin, myoglobin
 Toxic protein – ricin, trypsin inhibitor, lectins
 Structural protein – collagen, keratin
 Contractile protein – actin and myosin of muscle
 Secretory proteins – Fibroin
 Exotic proteins- antifreeze glycoproteins

4.  Simple
 Simple proteins yield on hydrolysis, only amino acids.
 Conjugated
 Simple proteins combined with some non-protein
substances known as prosthetic groups.
Derived
 Derived by partial to complete hydrolysis from the
simple or conjugated proteins by the action of acids,
alkalies or enzymes.
Classification based on Solubility and
Composition

5. Simple Proteins
Simple proteins are further subclassified based on their solubility and their
coagulability
 Albumins – Soluble in water and salt solutions
 Globulins – Sparingly soluble in water but soluble in salt solutions
 Prolamines – Soluble in 70 – 80% alcohol, insoluble in water
 Histones – Soluble in salt solutions
 Protamines - Soluble in water – not coagulated by heat
 Glutelins – Insoluble in all the above solutions, soluble in acid and base
 Scleroproteins – Insoluble in water or salt solutions
Classification based on Solubility and
Composition

6. Albumins
• Soluble in water and salt solutions
• Coagulated by heat
• Salting out by saturated ammonium sulphate
•They are deficient in glycine
• Ovalbumin, lactalbumin, serum albumin, legumelin
Globulins
• Sparingly soluble in water, soluble in salt solutions (NaCl)
• Coagulated by heat
• Salting out at half saturation by ammonium sulphate
• Deficient in methionine
• Serum globulin, vitellin, tuberin, legumin
Classification based on Solubility and
Composition

7. Prolamines
• Insoluble in water, and absolute alcohol, but soluble in 70 – 80% alcohol
• Rich in proline
• They are deficient in lysine
• Gliadin of wheat, zein of maize, hordein of barley
Glutelins
• Insoluble in water, soluble in dilute acids and alkalies
• Mostly found in plants
• Rich in glutamic acid
• glutenin of wheat, oryzenin of rice
Classification based on Solubility and
Composition

8. Histones
• Soluble in salt solutions
• Basic proteins
• They are not readily coagulated by heat
• Globin of hemoglobin and nucleoproteins
Protamines
• Soluble in water
• They are not coagulated by heat
• Basic due to large quantities of arginine
• Tyrosine and tryptophan are usually absent.
• They are found in asociation with nucleic acids in
the sperm cells of certain fish.
Classification based on Solubility and
Composition

9. Scleroproteins
• Insoluble in water, dil acids and alkali
• Highly stable
• Rich in Glycine, alanine and proline
• Keratin (hair, horn, nail, feather)
• collagen (bone, skin)
• Elastin (ligament)
Classification based on Solubility and
Composition

10. Conjugated proteins
Nucleoproteins
Lipoproteins
Mucoproteins
Phosphoproteins
Metalloproteins
Chromoproteins
Simple proteins combined with some non-protein substances known as
prosthetic groups. They are further classified based on the nature of
prosthetic groups.
Classification based on Solubility and
Composition

11. Nucleoproteins
• These are simple basic proteins (protamines or histones) in
combination with nucleic acids as prosthetic group.
• They are important constituents of nuclei and chromatin.
Lipoproteins
• These are proteins conjugated with lipids such as
triacylglycerols, phospholipids and cholesterol.
Glycoproteins
•These are composed of simple proteins in combination with
carbohydrates like mucopolysaccharides. Eg., Ovomucoid from
egg white
Classification based on Solubility and
Composition

12. Phosphoproteins
• These are proteins containing phosphoric acid.
• Phosphoric acid is linked to the hydroxyl group of certain amino
acids like serine. Eg., casein of milk
Metalloproteins
• These are metal binding proteins.
• Transferrin is a protein that binds to iron
• Cerruloplasmin binds to copper.
Chromoproteins
• Proteins containing coloured prosthetic groups
• Haemoglobin, flavoprotein, cytochromes.
Classification based on Solubility and
Composition

13. Primary derived proteins
• Derived from slight changes in protein molecule and its properties.
• Proteans, Metaproteins, Coagulated proteins
Secondary derived proteins
• These are formed by progressive hydrolysis of the peptide bonds of
proteins.
• Protein → Proteoses → Peptones → Peptides
Derived proteins
Classification based on Solubility and
Composition

14. Formation of peptide
Dipeptide
 Tripeptide contains three amino acids
 Peptides contain less than 50 amino acids
 More than 50 amino acids – Polypeptide
 Proteins contain one or more polypeptides

15.  Glutathione is a tripeptide
 Glutathione (γ-Glutamyl cysteinyl glycine)
 Reduced form is represented as GSH
 Takes part in redox reactions.
Glutathione
Glutathione

16.  To perform their biological function, proteins fold into a three dimensional structure
which is known as the native conformation.
 Due to the complexity of structure, protein structure is described by four levels of
organization
 Primary structure
 Secondary structure
 Tertiary structure
 Quaternary structure
Levels of protein structure
Myoglobin Haemoglobin

17.  Primary structure refers to the amino acid sequence of
the polypeptide chains.
 It refers to the number of amino acids and the order in
which they are covalently linked together. It also refers
to the location of disulphide bridges, if any.
 The bonds that stabilize primary structure are strong
covalent, peptide bonds.
 At one end of the chain there is a free amino group (N-
terminal) and at the other a free carboxyl group (C-
terminal).
 Numbering of amino acid residues always starts at the
N-terminal end.
Primary structure

18.  The important steps involved in determining primary structure are
i. Determination of number of polypeptide chains or subunits present in a protein.
ii. Separation of polypeptide chains if more than one is present.
iii. Determination of amino acid sequence of the subunits.
iv. Elucidation of the position of disulfide bonds, if any, between and within the
subunits.
Primary structure
Disulphide bond

19.  Refers to the repeating structures in a polypeptide.
 Secondary structures include alpha helix, beta pleated sheets, triple helix etc.
 Secondary structure is stabilized largely by hydrogen bonds formed between the atoms
present in the peptide bonds.
 The peptide bond exhibits partial double bond character. It is a rigid bond. The bonds
contributed by alpha carbon atom are free to rotate.
Secondary structure
α-Helix
β-Pleated
sheets
α-Helix

20. Alpha helix
Alpha Helix
 The amino acids in an α-helix are arranged in a right handed helical
structure.
 The peptide bonds form the backbone and side chains of amino acids
project outwards.
 The helix is stabilized by hydrogen bonds between the C=O group and the
N-H group of the first and fourth amino acids.
 There are 3.6 amino acid residues in one complete turn of the helix.
 The pitch of the helix is 5.4A
o
; Diameter of the helix is 5A
o
 Proline and glycine are called ‘helix breakers’ because they disrupt the α-
helical structure.
Proline
H2C
C=O
N-----
CH2
H2C
CH
Secondary structure

21. 21
 A β-strand is a stretch of amino acids whose peptide backbones are almost fully extended.
 They are sheet-like with zigzag conformation. Hence called pleated sheets.
 The beta strands are arranged side by side, which are cross-linked by hydrogen bonds.
 The polypeptide chains may run in parallel or antiparallel directions
 The strands become adjacent to each other, forming beta- sheet.
Beta pleated sheets
Secondary structure

22. Antiparallel
Parallel
 The polypeptide chains may run in parallel or antiparallel directions
 The strands become adjacent to each other, forming beta- sheet.
Beta pleated sheets
Secondary structure

23. Tertiarystructure
 Refers to the overall three dimensional conformation of a polypeptide chain.
 It indicates how the secondary structural features (helices, beta sheets, bends, loops)
assemble together forming a stable three dimensional structure.
 The proteins are folded in such a manner that the hydrophobic side chains are present
interior and hydrophilic side chains in the exterior of the molecule.

24.  The tertiary structure is stabilized by H-bonds, hydrophobic bonds, ionic
bonds (salt bridges), disulphide bridges, and weak van der Waals forces.
Tertiarystructure

25. Quaternary structure
 The way in which two or more individual folded polypeptide chains come together
and form a native conformation is described by the quaternary structure.
 This structure is applicable only to proteins having two or more polypeptide chains
(oligomeric proteins).
 The quaternary structure is mainly stabilized by weak bonds like hydrogen bonds
and also by disulphide bridges.
 Eg: Hemoglobin in RBC contains two alpha subunits and two beta subunits.

26. Brief on Structure of Protein

27. Protein folding
 Protein folding is the physical process by which a protein chain acquires its
native 3-dimensional structure.
 The amino-acid sequence of a protein determines its native conformation.
 A protein molecule folds spontaneously during or after biosynthesis.

28.  The process of folding often begins co-translationally, so that the N-terminus
of the protein begins to fold while the C-terminal portion of the protein is still
being synthesized by the ribosome.
 Specialized proteins called chaperones assist in the folding of other proteins.
 Although most globular proteins are able to assume their native state
unassisted, chaperone-assisted folding is often necessary in the crowded
intracellular environment to prevent aggregation.
 The passage of the folded state is mainly guided by hydrophobic interactions,
formation of intramolecular hydrogen bonds, ionic bonds and van der Waals
forces.
Protein folding

29. Physical and chemical properties
Physical properties
 Pure proteins are tasteless and odourless.
 Proteins exhibit colloidal properties.
 Proteins are amphoteric and possess both acidic and basic groups.
 They possess charged groups and migrate in electric field.
 The pH at which a protein exists as neutral molecule is called isoelectric pH (pI).
At isoelectric pH, a protein has least solubility and can be precipitated.
 Proteins are very sensitive to pH, UV radiation, heat and many organic solvents.
 Proteins have maximum absorption at 280nm due to the presence of tyrosine and
tryptophan. Hence the absorbance of the protein at this wavelength is used for its
determination.

30. Physical and chemical properties
Denaturation
 The phenomenon of altering the native conformation of protein is termed denaturation.
 During denaturation, the weak forces (Hydrogen bond, hydrophobic bond, ionic bond,
Van der waals force) responsible for maintaining secondary, tertiary and quaternary
structure of proteins are destroyed.
 The covalent peptide bonds are not broken. The primary structure is not affected.
 Denatured protein loses its function.
 Denaturation is caused by certain factors
 Phyical agents - heat, UV light, ultra sound, etc
 Chemical agents - Acid, alkali, heavy metal salts, urea, ethanol, etc
Native form (active) Denatured (inactive)
Denaturation
Renaturation

31. Physical and chemical properties
Denaturation
 Heat and UV radiation supply kinetic energy to protein molecules causing their atoms to
vibrate rapidly, thus disupting the weak hydrogen bonds and salt linkages. This results in
denaturation and coagulation of proteins.
 Organic solvents such as ethanol and acetone are capable of forming hydrogen bonds
with proteins. Hence addition of these solvents, will disrupt the hydrogen bonds present in
the secondary, tertiary and quaternary structures of proteins and cause denaturation and
precipitation.
 Acidic and basic reagents cause changes in pH, which alter the charges of side chains
that disrupt the salt linkages (ionic bond).
 Salts of heavy metals form very strong bonds with carboxylate ions of aspartate and
glutamate thus disturbing the salt linkages.
Renaturation
 Renaturation refers to the attainment of an original three dimensional functional protein
after denaturation.

32. Physical and chemical properties
Chemical properties
1. Biuret reaction
 Biuret test is highly used to detect presence of proteins in biological samples.
 Compounds containing two or more peptide bonds answer this test.
 This is due to the formation of coordination complex between four nitrogen atoms of two
polypeptide chains and one copper atom.
Cu2+
CH
CH NH
HN
R1 R3
CO
CO
R2
CH
NH CO
CH
CH NH
HN
R1
R3
CO
CO
R2
CH
NH CO

33. Physical and chemical properties
Chemical properties
2. Xanthoproteic reaction
 Addition of concentrated nitric acid to protein solution produces yellow colour
on heating. This is due to nitration of the phenyl rings of aromatic amino acids
forming nitro derivatives.
 The yellow stains on the skin caused by nitric acid is the result of
xanthoproteic reaction.
3. Hopkins-Cole reaction
 Indole ring of tryptophan reacts with glacial acetic acid in the presence of
concentrated sulphuric acid and forms a purple coloured product.
 Glacial acetic acid reacts with concentrated sulphuric acid and forms glyoxylic
acid, which in turn reacts with indole ring of tryptophan in presence of
sulphuric acid forming a purple coloured product.