PROTEIN CHEMISTRY - II

By Dr ANURAG YADAV

Proteins are the building blocks of body.

They are linear polymer made of amino acids sequence.

They may be monomeric protein with single chain or

Oligomeric with many polypeptide chain.

Abnormal in protein structure will lead to molecular disease with

profound alteration in metabolic function.

Proteins are made up of carbon, hydrogen, oxygen, nitrogen as major

& posphate, sulphur as minor component .

STRUCTURAL HIERARCHY

PRIMARY STRUCTURE OF PROTEIN

Formation of peptide bond.

Peptide nomenclature.

Geometry of polypeptide backbone.

Features of peptide bond.

primary structure of protein means the order of amino acids in the

polypeptide chain and the location of disulfide bonds, if any.

Primary structure denotes the number and sequence of amino

acids in the protein.

The higher level of organisation is decided by primary structure.

Each polypeptide chain has a unique amino acid sequence

decided by the genes.

The primary structure is maintained by the covalent bonds of

peptide linkage.

FORMATION OF PEPTIDE BOND

-Amino acids are linked by peptide bonds

-α carboxyl group of one amino acid reacts with α-amino group of

another amino acid to form peptide bond

Or called as CO-NH bridge.

Proteins are made up of polymerisation of amino acid through

peptide bond.

PEPTIDE NOMENCLATURE

by conventionally the peptide bond chain with free amino end

i.e N-terminal end on left side, & the free carboxyl end C-

terminal end at right.

Incidently the protein synthesis also begin from N-terminal.

GEOMETRY OF POLYPEPTIDE BACKBONE

Anatomy of peptide bond:

1. Is essentially is a planar.

in a dipeptide, two amino acids linked by peptide

where 6 atoms lie in same plane α-C & CO of first amino acid ,

NH & α-C of second amino acid.

Peptide bond is a partial double bond in character.

the normal bond length between

C-N = 1.49 A°

C=N = 1.27 A°

But the peptide bond C-N is 1.32 A°which is between these two

bonds.

Considerably double bond character of peptide linkage prevent

rotation around axis of this bond.

Hence a peptide bond is considered to be rigid.

Peptide bond is a polar covalent bond:

the sharing electron reside close to the oxygen conferring

negative charge over the oxygen & partial positive charge over

nitrogen.

thus the bond carries net no charge.

Two geometrical isomers are formed .

- in trans form = two α-C are on opposite side .

- in cis form = two α-C are on same side of plane.

All peptide bonds in protein exist as trans- form

Features of peptide bond can be summarized as

a. Bond is planar

b. It has partial double bond character.

c. It is RIGID & hence rotation around bond is restricted.

d. Peptide bond is POLAR covalent linkage.

e. All peptide bonds in protein are in trans- form

DISULPHIDE BOND

Disulfide are usually formed between two cysteine residue to

form cystine.

Both contribute to structural strength & stability of protein.

BONDS & ANGLES ADJACENT TO THE PEPTIDE

LINKAGE

Unlike rigid peptide bond which does not allow rotation of bonds

adjacent to it between

a. α-carbon & carbonyl group

b. amino group & α-carbon are flexible and purely single bond.

The free of rotation around these two bonds of each amino acid

enable protein to fold in different ways .

Rotations about these bonds is called as dihedral/ tortion angle &

measured between -180°to +180°

PHI(Φ) & PSI(Ψ) ANGLES

ANGLES

Not all conformation structure is possible with rotation of the

phi & psi angles.

Ramachandran showed with his plot , more than 75% of it are

not favourable/forbidden bcz of local steric clashes b/w

atoms.

INSULIN

Ex for primary structure

Sanger described structure 1955

β cells of pancreas

Hypoglycemic hormone

STRUCTURE OF INSULIN

Composed of 2 chains

A and B chain

A chain – glycine chain , 21 AA

B chain – phenyl alanine chain 30 AA

Intra and inter disulphide bonds

STRUCTURE

MATURATION OF INSULIN

proinsulin insulin

86 AA 51 AA

Species variation

A -8 , 9 , 10 . B– C terminal

Human insulin

Porcine insulin

Bovine insulin

HIGHER LEVEL OF PROTEIN STRUCTURE

They include :

Secondary

Tertiary

Quaternary structure of protein

CONFIGURATION & CONFORMATION

Configuration : refer to geometric relationship among given set

of atoms.

conversion to different configurational alternative into one

another is possible only by breaking & making covalent bond.

Conformation : refer to spatial relationship of every atom to all

other in three dimensional structure of protein.

the interconversion occur not by disruption of covalent

bond but rupture & reinstallation of relatively weak non-

covalent forces.

FORCES STABILISING HIGHER PROTEIN

STRUCTURE

Non-covalent : hydrogen bond.

hydrostatic bond.

ionic bond.

van der waals force.

Secondary structure is the steric relationship of amino acids

close to each other.

It denotes configurational relationship b/w residues which are

about 3-4 amino acid apart in linear sequence.

Stabilizing force: non-covalent forces (hydrogen bond, ionic

bond, hydrophobic and van der waals forces)

SECONDARY STRUCTURE OF PROTEIN:

Hydrogen bond :electrical attraction between hydrogen atom in

a polar bond in one molecule & oxygen/nitrogen atom in a

polar bond of another molecule/ within the same molecule.

hydrogen donor & hydrogen acceptor

Since AA can rotate around Φ & Ψ , peptide chain is flexible &

can be bent into number of conformation.

Polypeptide chain folded into

regular α-helix ,β-sheet.

irregular forms- turns & loops.

As peptide bonds are regular

interval along polypeptide ,

hydrogen bond b/w them tends

to force the chain into a coiled

conformation known as α-helix

Features of α-helix:

-most stable

-formed with lowest energy

-coiled structure with tightly

coiled polypeptide backbone

forming inner part of helix

with side chain extending

outwards from the central

axis

It can be either right handed/ left

handed .

All α-helices in protein are right

handed.

Covalent & many non-covalent

stablize α-helix.

Each turn is 5.4A° &

accommodate 3.6 AA residues

per turn of helix.

Thus AA spaced 3 or 4 residue

apart in sequence are spatially

closure to one another & each AA

form hydrogen bond with 4th AA in

linear sequence.

Formed when 2 or more polypeptides

line up side by side.

Individual polypeptide - β strand

Each β strand is fully extended.

They are stabilized by H bond b/w N-H

and carbonyl groups of adjacent

chains.

BETA PLEATED SHEET

2 types

Parallel Anti -Parallel

N C N

N N C

C

C

Features of β-pleated sheet:

- Second type of regular repetitive pattern

- Peptide backbone of these sheet are partly extended with

pleated appearance.

- Distance btwn AA along β-strand is 3.5A°( 1.5A° in α-helix)

- Side chain of adjacent AA orient opposite directions.

- Sheets are stabilized by extension hydrogen bond.

Anti-parallel sheet: hydrogen bonds b/w NH & CO group

connect each AA to single AA on an adjacent strand.

Eg; Silk fibroin

Parallel sheet: hydrogen bond connect each AA on one strand

with two different AA on adjacent strand.

Eg; Flavodoxin

Triple helix:

- structural proteins collagen is rich in

proline & hydroxy proline & hence it

cannot form α-helix & β-pleated

sheet , instead it forms a triple helix.

- Triple helix is stablized by same force

which stabilize the α-helix & β-

sheets.

LOOPS & TURNS IN SECONDARY STRUCTURE:

- Many proteins are globular in shape & require reversal in

direction of their direction of chain, reverse turn fulfill its need.

- Sometime loop like structure known as omega loops acts

linking material b/w regular α-helix & β-sheet pattern

OMEGA LOOP

Compact annular bend (Reverse turns of the peptide back

bone)

One or more loops join successive beta sheets and alpha helix

Present over the surface of the proteins (>60AA ) to avoid steric

hindrance

Each omega loop consists of 5 to 15 AA residue

R group densely crowding in the core of the protein

These loops contribute to functional site of the protein

SUPER SECONDARY STRUCTURES

(MOTIFS)

Beta barrel β-meander motif

beta-alpha-beta motif Greek key motif

Certain groupings of secondary structural elements are

called motifs.

TERTIARY STRUCTURE OF PROTEIN:

Denotes over all three dimensional

arrangement & inter-relationship of

various region/domains of single

polypeptide chain.

protein become fully functional only

when it is organised into tertiary level.

Stabilized by : non-covalent bonds.

More compact in organisation.

Non-polar region are burried in interior

portion & more polar arranged on to

surface.

DOMAINS:

Secondary & tertiary structures of large polypeptide are

organised into structurally connected but functionally

independent units known as domains.

They act as independent functional units when they binds to

their specific ligands.

The two-domain protein glyceraldehyde-

3-phosphate dehydrogenase.

QUATERNARY STRUCTURE OF PROTEIN:

Results when the protein consist of two or more polypeptide

chains held together by non-covalent forces

Not all proteins are organised at quaternary level.

Each individual polypeptide is called a subunit & the protein as

whole known as multimeric protein.

Sub units are held together by non covalent interactions.

Eg: hemoglobin have component as 2α 2 β.

Hydrogen bond :electrical attraction between hydrogen atom in

a polar bond in one molecule & oxygen/nitrogen atom in a

polar bond of another molecule/ within the same molecule.

hydrogen donor & hydrogen acceptor

Hydrophobic interaction:

-Occur when interatomic distance as low as 3 to 4 A°

-b/w hydrophobic side chain of non-polar AA that reside close

to each other in the interior of protein structure.

-This interaction is not bcz of any attraction b/w non-polar

groups , but due to property of water molecule surrounding

them which push them together resulting in hydrophobic

interaction.

Electrostatic interaction;

-Occur b/w oppositely charged groups such as COO- & NH3+ of

basic amino acid

or

-b/w amino terminal & carboxyl group of protein which remain

on surface of protein donot interact with other charged group

from protein bcz of high dielectric constant of water molecule

near by.

Van der waals interaction:

They are the weakest of non-covalent forces occuring over

extremely short distance.

The distance at which the attractive force b/w two atom is

maximal & repulsive force in minimal is termed as van der waal

contact distance which is sum of van der waal radii of two

atom.

They contribute for structural stability of protein bcz of cumulative

effect.