Structure and Properties of Peptides

Structure and Properties of Peptides

Learning Outcomes

• Recognize the chemical features of the peptide bond (amide bond) as a condensation reaction between two amino acids.

• Describe the chemical features of the peptide bond, including its resonance, planarity, and the orientation of the α-carbon of adjacent amino acid residues.

• Draw a polypeptide given the amino acid sequence (in single- and three-letter code).

• Determine the isoelectric point (pI) of a polypeptide given its amino acid sequence and the pH of the solution.

• Recognize and draw the titration curve for a given polypeptide.

• Predict if a polypeptide will be positive, negative, or zwitterionic at a given pH.

Peptide Bond Formation

Condensation Reaction Between Amino Acids

The peptide bond is formed by a condensation (dehydration synthesis) reaction between the α-carboxyl group of one amino acid and the α-amino group of another, resulting in the loss of a water molecule and the formation of a covalent amide linkage.

• General reaction: The carboxyl group (–COOH) of one amino acid reacts with the amino group (–NH2) of another.

• Product: A dipeptide and water ($\ce{H2O}$) are formed.

• Directionality: Peptides have an N-terminus (free amino group) and a C-terminus (free carboxyl group).

Equation:

$\ce{Amino ext{-}acid extsubscript{1} + Amino ext{-}acid extsubscript{2} ->[condensation] Dipeptide + H2O}$

Chemical Features of the Peptide Bond

Planarity and Resonance

The peptide bond exhibits unique chemical properties due to resonance between the carbonyl oxygen and the amide nitrogen, imparting partial double-bond character to the C–N bond.

• Planarity: Six atoms (Cα, C, O, N, H, Cα) are coplanar, restricting rotation around the peptide bond.

• Resonance: The peptide bond can be represented by two resonance structures, contributing to its rigidity and stability.

• Bond angles and lengths: The partial double-bond character shortens the C–N bond compared to a typical single bond.

Resonance structures:

$\ce{R-C(=O)-NH-R' <-> R-C(-O^-)=N^+-H-R'}$

Cis and Trans Conformations of the Peptide Bond

Conformational Isomerism

The peptide bond can exist in cis and trans conformations, defined by the relative positions of the α-carbons (Cα) on either side of the bond.

• Trans conformation: Cα atoms are on opposite sides of the peptide bond; this is the most common and energetically favored form in proteins.

• Cis conformation: Cα atoms are on the same side; less common due to steric hindrance.

• Proline exception: The amino acid proline is more likely to form cis peptide bonds than other amino acids, with a typical trans:cis ratio of about 4:1.

Peptide and Protein Nomenclature

Naming and Sequence Directionality

Peptides are named by listing amino acid residues from the N-terminus to the C-terminus. Each amino acid in a peptide chain is referred to as a residue.

• Naming convention: The suffix "-ine" or "-ate" is replaced with "-yl" for all residues except the C-terminal residue, which retains its full name. For glutamine (Gln), asparagine (Asn), and similar, the final "e" is dropped and replaced with "yl".

• Example: A tripeptide with the sequence serine–alanine–valine is named serylalanylvaline (or seryl-alanyl-valine).

• Sequence notation: Peptide sequences are written from N-terminus (left) to C-terminus (right).

Drawing Peptides

2D Representations and Backbone Structure

Peptides can be drawn in two dimensions by first sketching the repeating N–Cα–C=O backbone, then adding hydrogen atoms and side chains (R groups).

• Backbone: The backbone consists of repeating units of N–Cα–C=O.

• Side chains: Each Cα is bonded to a unique side chain (R group) specific to each amino acid.

• Hydrogens: Add hydrogen atoms to the nitrogen and Cα atoms as appropriate.

• Charge: Indicate formal charges on the N-terminus, C-terminus, and any ionizable side chains.

Example: The tripeptide ASV (Ala-Ser-Val) can be drawn with all atoms shown, or as a "chicken wire" diagram with stereochemistry indicated.

Classification of Peptides and Proteins

• Oligopeptide: Short chains of amino acids (typically fewer than 15 residues).

• Polypeptide: Chains longer than 15 amino acids.

• Protein: Long polypeptide chains that fold into specific three-dimensional structures.

Ionizable Groups and pKa Values in Proteins

pKa Ranges of Common Ionizable Groups

Proteins contain several ionizable groups, each with characteristic pKa values. These groups determine the overall charge of the peptide at a given pH.

Group	Typical pKa Range
α-Carboxyl	3.5–4.0
Side chain carboxyl (Asp, Glu)	4.0–4.8
Imidazole (His)	6.5–7.4
Sulfhydryl (Cys)	8.5–9.0
Phenolic (Tyr)	9.5–10.5
α-Amino	8.0–9.0
Side chain amino (Lys)	9.8–10.4
Guanidino (Arg)	~12

Isoelectric Point (pI) of Peptides

Calculation of pI

The isoelectric point (pI) is the pH at which a peptide or protein has a net charge of zero. It is determined by the pKa values of all ionizable groups in the molecule.

1. List all pKa values for the ionizable groups in the peptide.

2. Rank the pKa values from lowest to highest.

3. Draw the molecule at pH = 0 (all possible protons present) and determine the net charge (n).

4. Find the nth and (n+1)th pKa values in the ranked list.

5. Average these two pKa values to determine the pI: $pI = \frac{pK_{a,n} + pK_{a,n+1}}{2}$

Example: For the tetrapeptide EGAK (Glu-Gly-Ala-Lys), with pKa values 1.8, 4.2, 7.8, and 10.0, and net charge at pH 0 of +2, the pI is calculated as:

$pI = \frac{4.2 + 7.8}{2} = 6.0$

Titration Curves of Peptides and Proteins

Charge States and Isoelectric Point

Titration curves plot the net charge of a peptide or protein as a function of pH, showing the stepwise loss of protons from ionizable groups. The isoelectric point (pI) is the pH at which the net charge is zero.

• At pH below pI: The peptide is positively charged.

• At pH above pI: The peptide is negatively charged.

• At pH = pI: The peptide is zwitterionic (net charge = 0).

Application: Knowledge of pI is important for techniques such as ion exchange chromatography and protein purification.

Summary Table: Ionizable Groups and Their pKa Values

Ionizable Group	Typical pKa	Charge When Protonated	Charge When Deprotonated
α-Carboxyl	3.5–4.0	0	-1
α-Amino	8.0–9.0	+1	0
Asp/Glu side chain	4.0–4.8	0	-1
His side chain	6.5–7.4	+1	0
Cys side chain	8.5–9.0	0	-1
Tyr side chain	9.5–10.5	0	-1
Lys side chain	9.8–10.4	+1	0
Arg side chain	~12	+1	0

Additional info: The above notes expand on the provided slides by clarifying the process of pI calculation, the significance of peptide bond resonance, and the practical implications of peptide charge states in biochemical techniques.