Peptides

PeptidesHow are amino acids joined together in peptides and and proteins ?

A peptide bond may join two amino acids ————> dipeptide.

H2N HC

C

O

OH

CH3

HN

H

CH

HC

CH3CH3

C

O

OH

Alanine Valine

H2N HC

C

O

CH3

N

H

CH

HC

CH3CH3

C

O

OH

Alanylvaline (Ala-Val)

+ H2O

Peptide bond

Peptides

The Peptide Bond

The peptide bond is usually drawn as a single bond, but actually has considerable double bond character which prevents free

rotation about the bond:The atoms of the double bond, and those directly attached to it, all lie in the same plane.

The Peptide Bond

The trans-planar nature of the peptide bond accounts for the very high melting and boiling points and a lack of basicity in the simple

amides, and plays an important role in determining three-dimensional structure and function in polypeptides.

Drawing and Naming Peptides

1) Different constitutional isomers are possible whenever amino acids react to form peptides. (Ala-Val ≠ Val-Ala)

2) All peptides have one free α-amino group (N-terminal or amino-terminal residue), and one free α-carboxyl group (C-terminal or carboxyl-terminal residue).

3) When drawing (or naming) peptides the standard convention is to place the N-terminal residue at the left and the C-terminal residue to the right.

Drawing a Peptide

H2N HC

C

O

OH

R

HN

H

CH

R'

C

O

OH

H2N HC

C

O

R

H

NCH

R'

C

O

OH

H2N HC

C

O

R H

NCH

R'

C

O

OH

Peptide bond forms here

Peptide bond

H2O removed through

condensation

Ionization of Peptides1) As in the case of amino acids, each peptide has an isoelectric pH (or “pI”) at which it does not migrate in an electric field.

2) The pI value of a peptide containing only neutral amino acid residues or equal numbers of acidic and basic residues or both is in the range of pI values for neutral amino acids (pH 5.05–6.30).

3) The pI of a peptide containing acidic and basic amino acid residues is

a) on the acidic side (lower than 5.05–6.30) if there is an excess of acidic residues, and

b) on the basic side (higher than 5.05–6.30) if there is an excess of basic residues.

4) All amino and carboxyl groups, including those on side groups of acidic and basic amino acid residues, are charged at physiological pH.

Physical Properties of Peptides

1) Peptides have solubility and electrophoresis properties that are pH dependent.

2) Peptide solubility is lowest at its isoelectric point.

3) At a given pH, each peptide has a particular electrical charge depending upon its isoelectric point and the number of ionizable groups it contains.

4) Electrophoresis can be used to separate peptides of differing charges

5) Peptides are often distinguished from proteins by the number of amino acid residues. Molecules having fewer than 50 amino acid residues are generally called peptides, regardless of physiological activity.

Chemical Properties of Peptides

HN

HC C

O

CH2

SH

NH

CH

C

O

CH2

SH

NH

CH

C

O

CH2

S

HN

HC C

O

CH2

S

peptide chain

peptide chain

reduction

oxidation

two cysteine residues

cystine disulfide bridge

Pairs of cysteine residues often link two peptide chains or two parts of one peptide chain through disulfide bridges:

Chemical Properties of PeptidesDisulfide bridges in peptides may be represented using the

3 letter amino acid abbreviations.

Disulfide bridges generally survive hydrolysis reactions, so a separate chemical reaction is often necessary to isolate the amino

acids involved.

Examples of Physiologically Active Peptides

Aspartame - 2 amino acids An Artificial Sweetener

Met enkephalin - 5 amino acids Reduces Pain Sensation

Bradykinin - 9 amino acids Powerful Vasodilator-Released by mast cells after injuries and

during allergic responses. Similar to histamine in actions

Oxytocin 9 amino acids; 1 disulfide linkage

Causes contraction of uterine muscles during labor.

Insulin 51 amino acids ; 3 disulfide linkages

Regulator of carbohydrate metabolism and absorption

Insulin 51 amino acids ; 3 disulfide linkages

Regulator of carbohydrate metabolism and absorption

Insulin Maturation

Protein Structure and

Function

Classification of Proteins Based on Components

Protein Structure

Simple proteins - Proteins containing only polypeptides

Conjugated proteins - Proteins containing nonpolypeptide molecules or ions

1) Apoprotein - The polypeptide part of a conjugated protein

2) prosthetic group - The nonpolypeptide part of a conjugated protein

Protein Structure

Classification of Conjugated Proteins

1) Glycoproteins

2) Hemoproteins

3) Lipoproteins

4) Metalloproteins

5) Nucleoproteins

6) Phosphoproteins

Three-Dimensional Structure of Proteins

The conformations of the individual bonds in all the amino acid residues within the protein produces a unique 3-D shape, which in turn produces a unique physiological function.

The overall folding of a protein is described at four levels:

1º 2º 3º 4º

Levels of Protein Structure

• Primary structure is the amino acid sequence of a polypeptide.

• Secondary structure is the conformation in a local region of a polypeptide molecule. The conformation usually involves a regular coiling or layering of the protein chain.

• Tertiary structure exists when the polypeptide has different secondary structures in different local regions. Tertiary structure describes the three-dimensional relation among the different secondary structures in different regions.

• Quaternary structure exists only in proteins in which two or more polypeptide molecules aggregate together. It describes the three-dimensional relationship between the different polypeptides.

The most stable conformation of a protein is determined by:

1) The bonds in the linear chain

a) No free rotation about the peptide bond . b) Limited rotations about the bonds of the alpha carbon

2) Hydrogen-bonding between peptide amide bonds from different residues

3) Interactions of side chains with each other & with water

a) The tendency of non-polar side chains to avoid water

b) The attraction of non-polar side chains for each other

c) Hydrogen bonding between polar side chains & water

d) Ionic attractions between charged side chains

e) Disulfide bonds between side chains

Hydrophobic effect

Determinants of Protein Conformation

1º

2º3º

Secondary Protein Structure - The α-Helix

The polypeptide chain is arranged like a coiled spring with a hydrogen bond between each peptide group’s C=O oxygen and the hydrogen of the N-H group of the fourth residue farther down the chain.

Secondary Protein Structure - The β-Pleated Sheet

Peptide chains are extended and run side-by-side each other in either a parallel or an antiparallel arrangement. Neighboring chains are held together by hydrogen bonds between an N-H on one chain and a C=O on a neighboring chain. Side chains extend alternately above and below the plain of the sheet.

Secondary Protein Structure - The β-Pleated Sheet

Levels of Protein Structure

Levels of Protein Structure

Representation of a Simple Protein

Protein StructureStructural Classification of Proteins

Fibrous Proteins

Silks, Keratins, Collagens

Globular Proteins

Enzymes, Antibodies, Hormones

1) Elongated, water insoluble

2) Structural and contractile functions

3) No tertiary structure, but generally possess a single conformational pattern throughout most of the chain (secondary structure)

4) Most have a quaternary structure involving the aggregation of polypeptide chains

1) Remain soluble in water in order to carry out their metabolic functions

2) Spherical, Globular

3) Remain water soluble by folding up so as to segregate hydrophobic side chains in the interior of the molecule and hydrophilic side chains on the exterior of the molecule

Fibrous Proteins

α-Keratins

The structural component of hair, horn, hoofs, nails, skin, and wool.

These materials have a hierarchical structure. Coiling at higher and higher levels is a mechanism for enhancing physical strength.

Fibrous Proteins

α-Keratins

The packing within the α-keratins is stabilized by disulfide bridges and secondary forces between different polypeptide

molecules.

Disulfide bridges are more important than secondary forces in imparting insolubility, strength, and resistance to

stretching.

Interchain disulfide bonds are often called cross-links.

The degree of hardness of an α-keratin depends upon its degree of cross-linking. High cysteine content results in

increased hardness (hair, horn, nail) compared to low cysteine content (skin, callus).

Fibrous Proteins“Permanent” Hair Waving

Permanent waving of hair is accomplished by breaking and reforming cysteine cross-links within the hair fiber:

β-Keratins and Silk Fibroins

The β-keratins make up the proteins in bird feathers, reptile scales, and silk fibroin.

β-Keratins are almost completely composed of β-pleated sheets.

Fibrous Proteinsβ-Keratins

Fibrous Proteins - Collagen

The most abundant protein in vertebrates is collagen.

Collagen is a stress-bearing component of connective tissues such as bone, cartilage, cornea, ligament, teeth, tendons, and the fibrous matrices of skin and blood vessels.

Collagen contains much more glycine and proline and much less cysteine than does α-keratin. Much of the proline present is converted into hydroxyproline.

A single collagen molecule forms a left-handed helical structure, much more elongated than an α-helix. Three left-handed collagen helices twist around each other to form a right-handed superhelix called a triple-helix or tropocollagen.

Tropocollagen is further organized into fibrils and higher-level structures.

Fibrous Proteins - Collagen

0.5-3μm

10-300 nm

1.5 nm

diameter

Fibrous Proteins - Collagen

Globular Proteins

Globular proteins remain soluble by folding up in such a way as to segregate their hydrophobic amino acid side chains in the interior of the molecule, and their

hydrophilic amino acid side chains on the exterior of the molecule, in contact with water.

Globular proteins do not aggregate into macroscopic structures but remain soluble in order to carry out their

metabolic functions:

catalysis, transport, regulation, and protection.

Globular Proteins

Myoglobin and Hemoglobin

Hemoglobin in red blood cells binds oxygen in the lungs, transports it through the blood stream,

and releases it in the tissues.

Myoglobin has a higher affinity for oxygen than does hemoglobin and is found in muscle tissue.

Myoglobin serves as a storage reserve for oxygen within the muscle.

Globular Proteins - Myoglobin

Myoglobin consists of a single polypeptide chain containing 153 amino acid residues, organized into 8 α-helical regions that surround a prosthetic group

called a heme group.

The iron atom on the heme group is the site of attachment of the O2 molecule.

Heme Group

In 1971, Professor of Biochemistry F. R. Gurd assembled a model of myoglobin. The model was constructed of precisely bent wire segments to represent the amino acids, each fragment fastened to its neighbors using links that were screwed together. Wires were stretched throughout the structure to ensure that the various parts were held in proper alignment. A separate "space-filling" model is visible in the background. Prof. Gurd required three weeks and an entire 20 X 30 foot room to assemble the model. Today, a comparable three-dimensional model of myoglobin can be displayed in a matter of seconds. (Univ of Indiana)

Levels of Protein Structure

Globular Proteins - Hemoglobin

4 polypeptide chains 2 α-chains (141 residues each) 2 β-chains (146 residues each) Each α and β chain folded in a manner similar to that of myoglobin contains a heme group capable of carrying oxygen

HEMOGLOBIN A GLOBULAR PROTEIN WITH

QUATERNARY STRUCTURE

The surfaces of the α and β chains contain some hydrophobic residues which cause all 4 chains to aggregate into a tetramer. A space at the center of the tetrameric structure can bind a molecule of 2,3-bisphosphoglycerate (BPG) which regulates the affinity of the hemoglobin molecule for oxygen.

Globular Proteins - Denaturation

Denaturation - the loss of native conformation due to a change in environmental conditions. The non-functioning protein is called a denatured protein.

Denaturation results from the disruptions of the weak secondary forces holding a protein in its native conformation. (Disulfide bridges confer considerable resistance to denaturation because they are much stronger than the weak secondary forces.)

Globular Proteins - Denaturation

A variety of denaturing conditions or agents lead to protein denaturation:

1) Increased temperature (or microwave radiation)

2) Ultraviolet and ionizing radiation

3) Mechanical energy

4) Changes in pH

5) Organic chemicals

6) Heavy metal salts

7) Oxidizing and reducing agents

The sickle cell mutation causes hemoglobin molecules to clump t o g e t h e r i n a n a b n o r m a l manner. The valine in position 6 adheres to a notch on the oppos i te s ide o f another molecule of Hb, causing long chains to form.

In sickle cell anemia, the normal hemoglobin molecule mutates by exchanging the 6th amino acid on the beta chain from glutamic acid to valine. Normal Hb has the genotype SS. Sickle cell anemia occurs when an individual inherits two recessive alleles (ss). Sickle cell trait exists when one inherits the heterozygous condition (Ss). The malaria parasite (Plasmodium falciparum) does not survive in these individuals; they may have a slight anemia, but they survive better than either normal individuals (SS- who often die of malaria), or those who die of sickle cell disease (ss).

Sickle Cell Anemia

Comparison of the distribution of malaria (left) and sickle-cell anaemia (right) in Africa

Sickle Cell Anemia