25.

15
Peptide Bond Formation
 Peptide Bond Formation

To form a peptide bond between two suitably protected amino

acids, the free carboxyl group of one of them must be
activated so that it is a reactive acylating agent.

N,N-Dicyclohexylcarbodiimide (DCCI)has the structure:

 Mechanism of Peptide Bond Formation

Reaction equation.
 Mechanism of Peptide Bond Formation

Step 1. Deprotonation, nucleophilic attack.

Step 2. Nucleophilic attack, proton transfers.

 Mechanism of Peptide Bond Formation

Step 3. Dissociation.
 Peptide Bond Formation

Higher peptides are prepared either by:

(a) stepwise extension of peptide chains, one amino acid

at a time, or by:

(b) coupling of fragments containing several residues

(the fragment condensation approach).

Either way the solid phase method is superior.

 25.16
Solid-Phase Peptide Synthesis:
The Merrifield Method
Solid-Phase Peptide Synthesis

Step 1: The Boc-protected amino

acid is anchored to the resin.

Step 2: The Boc protecting group

is removed by treatment with HCl
in dilute acetic acid.

Step 3: The resin-bound C-terminal

amino acid is coupled to an N-
protected amino acid
Solid-Phase Peptide Synthesis

Step 4: The Boc protecting

group is removed.

Step n: The peptide is

removed from the resin by
treatment with HBr in
trifluoroacetic acid.
 Solid-Phase Peptide Synthesis

Yields of 99% and greater are achieved with current

instrumentation.
Thousands of peptides and peptide analogs have been
prepared by the solid-phase method.

A section of polystyrene modified by chloromethylation.

 25.17
Secondary Structures of
Peptides and Proteins
Secondary Structures of Peptides and Proteins

The secondary structure is the conformational relationship

of nearest neighbor amino acids with respect to each other.

Two arrangements, the a helix and the b sheet are

secondary structural units that are both stable and
common.

Two important features:

1. The peptide bond is planar and the main chain is in an
anti conformation.
2. Hydrogen bonding occurs when the NH of one amino
acid and the C=O of another are close in space;
conformations with more H-bonds are stabilized.
Secondary Structures of Peptides and Proteins

Both the parallel and antiparallel arrangements of chains

occur in proteins.
Secondary Structures of Peptides and Proteins

The b-pleated sheet secondary structure of a protein,

composed of alternating glycine and alanine residues.
Secondary Structures of Peptides and Proteins

The a helix is another commonly encountered secondary

structure.
Secondary Structures of Peptides and Proteins

Molecular models of ribonuclease. Red ribbons identify

sequences where the secondary structure is a helix; yellow
ribbons identify strands of sheet.
Secondary Structures of Peptides and Proteins

Barrel-shaped green fluorescent protein (GFP) has an outer

b-sheet structure and an a helix in the inner region.
 25.18
Tertiary Structure of
Polypeptides and Proteins
 Tertiary Structure of Polypeptides and Proteins

The way a protein chain is folded, its tertiary structure, affects

its physical properties and its biological function.

The two main categories of tertiary structure are:

1. Fibrous proteins. Bundles of elongated filaments of

protein chains that are insoluble in water.

2. Globular proteins. Approximately spherical and are either

soluble or form colloidal dispersions in water.
Tertiary Structure of Polypeptides and Proteins

Two a helices combine to give a coiled coil and a pair of

coiled coils form a protofilament. Four protofilaments give
a filament - the structural material from which the fibrous
protein is assembled.
 Tertiary Structure of Polypeptides and Proteins

The structure of carboxypeptidase A.

The tube model (left) emphasizes the globular shape.
The ribbon diagram (right) emphasizes the folding of the chain.
 Carboxypeptidase-Catalyzed Hydrolysis

The mechanism shown outlines the major stages in carboxy-

peptidase-catalyzed hydrolysis of a peptide in which the C-
terminal amino acid is phenylalanine.

Stage 1.
 Carboxypeptidase-Catalyzed Hydrolysis

Stage 2.

Stage 3.
 25.19
Coenzymes
 Coenzymes

Proteins act with substances known as cofactors.

Cofactors may be organic or inorganic and strongly or

weakly bound to the enzyme.

The term coenzyme is applied to those that are not

covalently bound to the enzyme, and prosthetic group to
those that are.

Heme is an important prosthetic group

 Coenzymes

Heme has a coplanar arrangement of the groups

surrounding iron.
 Coenzymes

Heme in sperm-whale myoglobin.

 25.20
Protein Quaternary Structure:
Hemoglobin
 Protein Quaternary Structure: Hemoglobin

Some proteins are assemblies of two or more chains. The

manner in which these subunits are organized is called the
quaternary structure of the protein.

Hemoglobin is the oxygen-carrying protein of blood.

Hemoglobin is an assembly of four hemes and four protein

chains, including two identical chains called the alpha chains
and two identical chains called the beta chains.
 Oh NO! Its Inorganic!

The amino acid L-arginine undergoes an interesting bio-

chemical conversion.

L-Arginine L-Citrulline Nitric oxide

Identifying NO as a signaling molecule in biological

processes clearly justified a Nobel Prize (1998).
 25.21
G-Coupled Protein Receptors
 G-Coupled Protein Receptors

G-coupled protein receptors, or GCPRs. The G stands for

guanine in guanine nucleotide-binding proteins.

GCPRs occur throughout the body and function as molecular

switches that regulate many physiological processes.

GCPRs are involved in many diseases. It is estimated that

nearly one-half of prescription drugs target GCPRs in the
treatment of cancer, cardiac malfunction, inflammation, pain,
and disorders of the central nervous system.
 G-Coupled Protein Receptors

Signal transduction is initiated by the binding of a G-coupled

protein receptor (GCPR) to the ligand on the exterior of the
cell.