Amino Acids, Peptides, and Proteins: Structure, Properties, and Biochemical Techniques

3.1 Amino Acids

Principles and Overview

Amino acids are the fundamental building blocks of proteins, and all living organisms use a common set of 20 amino acids. Each amino acid contains a side chain (R group) that imparts unique chemical properties, making them the 'alphabet' of protein structure.

• Key Point: Proteins are constructed from 20 standard amino acids.

• Key Point: The side chain (R group) determines the chemical nature of each amino acid.

• Example: Glycine is the simplest amino acid, with a hydrogen atom as its R group.

Common Structural Features

• Each amino acid has a central α-carbon bonded to four different groups:

  • A carboxyl group (–COOH)

  • An amino group (–NH2)

  • A hydrogen atom

  • An R group (side chain, unique to each amino acid)

• The α-carbon is a chiral center (except in glycine).

• The structure is tetrahedral.

Stereochemistry of Amino Acids

• Amino acids (except glycine) exist as two enantiomers: L and D forms.

• Proteins in nature are composed exclusively of L-amino acids.

• Enantiomers are optically active and the D, L system specifies absolute configuration.

Classification by R Group

Amino acids are classified into five main groups based on the properties of their R groups:

• Nonpolar, aliphatic (7)

• Aromatic (3)

• Polar, uncharged (5)

• Positively charged (3)

• Negatively charged (2)

Table: Amino Acid Classes and Examples

Class	Examples	Key Properties
Nonpolar, aliphatic	Glycine, Alanine, Proline, Valine, Leucine, Isoleucine, Methionine	Hydrophobic, stabilize protein structure
Aromatic	Phenylalanine, Tyrosine, Tryptophan	Absorb UV light (270–280 nm), hydrophobic effect
Polar, uncharged	Serine, Threonine, Cysteine, Asparagine, Glutamine	Form hydrogen bonds, cysteine forms disulfide bonds
Positively charged	Lysine, Arginine, Histidine	Significant positive charge at pH 7.0
Negatively charged	Aspartate, Glutamate	Net negative charge at pH 7.0

Uncommon Amino Acids

• Some amino acids are modified after protein synthesis (e.g., 4-hydroxyproline in collagen).

• Others are modified during synthesis (e.g., pyrrolysine in methane biosynthesis).

• Transient modifications (e.g., phosphorylation) regulate protein function.

• Some are free metabolites (e.g., ornithine in arginine biosynthesis).

Acid-Base Properties

• Amino acids can act as acids or bases due to their amino, carboxyl, and ionizable R groups.

• At neutral pH, amino acids exist as zwitterions (both positive and negative charges).

Titration and Buffering

• The carboxyl group has an acidic $pK_a$ (typically ~2.0–2.5).

• The amino group has a basic $pK_a$ (typically ~9.0–10.0).

• The isoelectric point (pI) is the pH at which the net charge is zero:

$pI = \frac{pK_1 + pK_2}{2}$

• Amino acids act as buffers near their $pK_a$ values.

• Ionizable side chains add complexity to titration curves and buffering capacity.

3.2 Peptides and Proteins

Peptide Bond and Structure

Amino acids are linked by peptide bonds (amide linkages) formed via condensation reactions and broken by hydrolysis. The sequence of amino acids in a protein is its primary structure.

• Dipeptide: 2 amino acids, 1 peptide bond

• Tripeptide: 3 amino acids, 2 peptide bonds

• Oligopeptide: A few amino acids

• Polypeptide: Many amino acids, MW < 10 kDa

• Protein: Thousands of amino acids, MW > 10 kDa

Peptide Terminals and Naming

• Numbering and naming start from the amino-terminal (N-terminal) residue.

• Peptides can be named using:

  • Full names (e.g., seryl-glycyl-tyrosyl-alanyl-leucine)

  • Three-letter codes (e.g., Ser-Gly-Tyr-Ala-Leu)

  • One-letter codes (e.g., SGYAL)

Ionization Behavior

• Peptides have ionizable groups: one free α-amino, one free α-carboxyl, and some R groups.

• This affects their charge and behavior in different pH environments.

Protein Size and Composition

• Proteins vary greatly in length and composition, from a few to thousands of amino acids.

• Multisubunit proteins contain two or more polypeptides; oligomeric proteins have at least two identical subunits (protomers).

• Number of residues can be estimated: Number of residues = Molecular weight / 110

Conjugated Proteins

• Some proteins contain non-amino acid components called prosthetic groups (e.g., heme, lipids, sugars, metals).

• Examples: Lipoproteins (lipids), glycoproteins (sugars), metalloproteins (metals).

3.3 Working with Proteins

Protein Purification and Separation

Proteins can be separated and purified based on size, charge, binding properties, and solubility. Purification is essential for studying protein function and structure.

• Step 1: Break open cells to obtain a crude extract containing proteins.

• Step 2: Fractionation separates proteins by size or charge (e.g., salting out).

• Step 3: Dialysis uses a semipermeable membrane to separate proteins from small solutes.

Chromatography Techniques

• Column Chromatography: Proteins migrate through a solid phase; migration depends on protein properties.

• Ion-Exchange Chromatography: Separates proteins by net charge using cation or anion exchangers; pH and salt concentration affect separation.

• Size-Exclusion (Gel Filtration) Chromatography: Separates by size; larger proteins elute first.

• Affinity Chromatography: Separates by specific binding affinity; elution by high salt or ligand concentration.

• High-Performance Liquid Chromatography (HPLC): Uses high pressure for improved resolution.

Tables

Table: Representative Amino Acid Properties (from Table 3-1)

Amino Acid	3-Letter Code	1-Letter Code	pKa (COOH)	pKa (NH3+)	pKa (R group)
Glycine	Gly	G	2.34	9.60	-
Glutamate	Glu	E	2.19	9.67	4.25
Lysine	Lys	K	2.18	8.95	10.53
Histidine	His	H	1.82	9.17	6.00
Tyrosine	Tyr	Y	2.20	9.11	10.07

Key Equations

• Isoelectric Point (pI): $pI = \frac{pK_1 + pK_2}{2}$

• Number of Residues: $\text{Number of residues} = \frac{\text{Molecular weight}}{110}$

Summary

• Amino acids are the monomers of proteins, classified by their R groups and chemical properties.

• Peptide bonds link amino acids into peptides and proteins, with structure and function determined by sequence.

• Proteins can be purified and analyzed using a variety of biochemical techniques, including chromatography and electrophoresis.

• Understanding amino acid properties and protein purification is foundational for advanced study in biochemistry.