The peptide bond forms spontaneously via a condensation reaction that releases a small molecule like water. It has partial double bond character that restricts rotation, resulting in planar geometry. The phi and psi angles of the peptide backbone determine its conformation within allowed regions of the Ramachandran plot, avoiding steric clashes. While most peptide bonds adopt the trans configuration, proline favors cis. Protein function arises from its precise three-dimensional structure.

1. Function Arises from Conformation: The Peptide Bond

2. Peptide bonds
What is the mechanism of peptide bond formation?

3.  The process is spontaneous and known as a condensation reaction or dehydration
reaction.
 It is a chemical reaction in which two molecules or moieties (functional groups) combine
to form a larger molecule, together with the loss of a small molecule.
 Possible small molecules lost are water, hydrogen chloride, methanol, or acetic acid.
Is this reaction spontaneous or requires any help?

4. How much time does this reaction require to complete the process of the product formation?
The enzyme unease catalyzes the reaction of urea with water to produce carbon dioxide and
ammonia with at least 104 to 105 fold higher than unanalyzed reaction.

5. Peptide bond formation on the ribosome
About 20% of the cellular energy is used for making peptide bonds.

6. The proposed mechanism of peptide bond formation on the ribosome
Hiller et al., 2011, nature, 476, 236-240.

7. Properties of peptide bonds
The peptide bond formed between the carboxyl and amino groups of two amino acids is a
unique bond that possesses little intrinsic mobility. This occurs because of the partial double
bond character.
On average , a peptide bond length is 1.32 Å compared to 1.45 Å for an ordinary C-N bond.
In comparison the average bond length associated with a C=N double bond is 1.25 Å.
Thus, partial double bond restricts rotation about this bond. This leads to the six atoms being
coplanar.
Ca
Ca

8. Can accept and donate H-bonds (the peptide bond is not inert).
Possesses a dipole: the H in NH is positively charged and the O in CO is negatively charged.

9. Cis/trans isomers of the peptide group
In the unfolded state of proteins, the peptide groups are free to isomerize and adopt both
isomers; however, in the folded state, only a single isomer is adopted at each position.
For most peptide bonds, the ratio of cis to trans configurations is approximately 1:1000
(except for proline where it is 1:4 ratio).
As a result of restricted motion about the peptide bond two conformations related by an angle
of 180 are possible (Ca atoms in Trans and Cis with respect to peptide bond).

10. A peptide bond can be broken by hydrolysis (the adding of water).
Peptide bond hydrolysis
This process is extremely slow (up to 1000 years).
Proteases catalyze amide (peptide) bond hydrolysis in protein or peptide substrates:
Peptide bond hydrolysis is reversible or irreversible?
In the presence of water they will break down and release 8–16 kJ/mol (2–4 kcal/mol) of free
energy. The equilibrium of this reaction lies on the left side. I.e. hydrolysis is prefered to
synthesis.

11. Torsion angles: Phi & Psi
• Rotational constraints emerge from interactions with bulky groups (i.e. side chains).
• The dihedral angles at Ca atom of every residue provide polypeptides requisite
conformational diversity, whereby the polypeptide chain can fold into a globular shape.
For any polypeptide backbone represented by the sequence -N-Cα-C-N-Cα-C-, only the N-Cα
and Cα-C bonds exhibit rotational mobility.

12. Proteins: polymers of amino acids
When joined in a series of peptide bonds amino acids are
called residues to distinguish between the free form and the
form found in proteins.
Cα atoms
Backbone atoms
Main chain atoms

13. Thereby the peptide has a direction: the N-terminus is the start and the C-terminus is the end.
Total number of protein sequences of length L is equal to 20L. However, not all amino acids
are found in equal frequency in proteins.

14. Most natural polypeptide chains contain between 50 and 2000 amino acid residues and are
commonly referred to as proteins.
Peptides made of small numbers of amino acids are called oligopeptides or simply peptides.
The mean molecular weight of an amino acid residue is about 110, and so the molecular
weights of most proteins are between 5500 and 220,000.

15. Proteins can only have a function if they have the correct conformation for it (function arises
from conformation).
Polypeptide Chains Are Flexible Yet Conformationally Restricted

16. There is rotation around the bonds between N-Cα and Cα-C, but these allow only rotation of
the Cα (and the other atoms linked to them).
Rotating the Cα does not move it outside the plane (only the atoms linked to it are moved).
A fully extended polypeptide chain has Φ = Ψ = 180 .

17. Amino acids with a side chain of more than one atom can also have a rotation at the Cβ. But
only a certain number of conformations is allowed.
Conformers which differ only by rotation about a single bond are termed rotamers. Usually a
staggered conformation is preferred.
Side chain mobility

18. In polypeptide chemistry the term "conformation" should be used, in conformity with current
usage, to describe different spatial arrangements of atoms produced by rotation about covalent
bonds; a change in conformation does not involve the breaking of chemical bonds (except
hydrogen bonds) or changes in chirality.
On the other hand in polypeptide chemistry the term "configuration" is currently used to
describe spatial arrangements of atoms whose interconversion requires the formal breaking
and making of covalent bonds.
Conformation and Configuration

19. Ramachandran Plot
Are all combinations of φ and ψ possible?
G. N. Ramachandran recognized that many combinations are forbidden because of steric
collisions between atoms. The allowed values can be visualized on a two-dimensional plot
called a Ramachandran diagram.

20. Three-quarters of the possible (φ and ψ) combinations are excluded simply by local steric
clashes. Steric exclusion, the fact that two atoms cannot be in the same place at the same time,
can be a powerful organizing principle.

21. Assignment:
Sketch the Ramachandran
plot between 0-360 phi and
psi angles.

22. Thermus aquaticus EFTu-GDP: Arg345, Arg274
and Leu258 lie in disallowed regions.
Arg345 is in position 2 of a type I reverse turn.
No other conformation of Arg345 would allow the
end of its side chain to reach the surface of the
domain.
Arg274 is in position 3 of a type II reverse turn.
Leu258 is in a loop connecting 2 β-strands. (The
structure of the protein in a crystal might be
different from that in its normal environment. So
some outliers may be caused by crushing the
protein to fit in the crystal structure.
What if there are residues in the disallowed region of the Ramachandran Plot?

23. Arg90
Thymidylate Kinase (2PBR)

24. Methods of peptide conformation studies
 Nuclear Magnetic Resonance (NMR)
 Hydrogen Exchange
 Fluorescence Resonance Energy Transfer (FRET)
 Circular Dichroism (CD)