Protein Three-Dimensional Structure: Primary and Secondary Structures

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Protein Three-Dimensional Structure

Introduction to Protein Structure and Function

Proteins are essential biological macromolecules whose function is determined by their three-dimensional (3D) structure. The unique folding of a polypeptide chain into a specific 3D conformation under physiological conditions is critical for its biological activity.

Native conformation: The functional, folded form of a protein under normal conditions.
Structure-function relationship: The specific 3D structure of a protein enables its unique biological function; even small changes can disrupt activity.
Examples: Hemoglobin, catalase, myosin, DNA-binding proteins, and enzymes all rely on precise 3D structures.

Levels of Protein Structure

Hierarchy of Protein Structure

Proteins exhibit four levels of structural organization, each contributing to the overall shape and function of the molecule.

Primary structure: The linear sequence of amino acids (aa) in a polypeptide chain.
Secondary structure: Regular, recurring local conformations stabilized by hydrogen bonds, such as α-helix and β-sheet.
Tertiary structure: The overall 3D folding of a single polypeptide chain, including interactions between secondary structure elements.
Quaternary structure: The association of multiple polypeptide subunits into a functional protein complex.

Figure: Diagram showing primary, secondary, and tertiary structure organization. (Additional info: Quaternary structure is not always present; it occurs in proteins with more than one polypeptide chain.)

Peptide Bond Formation and Protein Backbone

Peptide Bond: Linking Amino Acids

Amino acids are joined by peptide bonds, which are covalent linkages formed between the α-carboxyl group of one amino acid and the α-amino group of another. This reaction is a condensation (dehydration) reaction, releasing water.

Peptide bond: A secondary amide bond; formation eliminates some ionizable groups.
Reaction equation:

Backbone structure: The repeating sequence of N–Cα–C forms the protein backbone, which has a high capacity for hydrogen bonding (via N–H and C=O groups).
Protein size: Proteins typically contain 50–2000 amino acids, with an average mass of ~110 g/mol per amino acid residue.

Amino Acid Residues and Directionality

Once incorporated into a peptide, amino acids are referred to as residues. The peptide chain has directionality, with the N-terminus (amino end) at one end and the C-terminus (carboxyl end) at the other.

Residue: An amino acid unit within a polypeptide chain.
Directionality: Chains are always written and synthesized from N-terminus to C-terminus.
Example: The pentapeptide sequence Tyr-Gly-Gly-Phe-Leu is written as N-YGGFL-C.

Covalent Cross-Linking: Disulfide Bonds

Disulfide Bridges in Protein Structure

Some proteins contain covalent cross-links called disulfide bonds, which form between the thiol groups of two cysteine residues. These bonds stabilize protein structure, especially in extracellular proteins.

Formation: Oxidation of two cysteine side chains forms a disulfide bond (cystine).
Reaction equation:

Reduction: Disulfide bonds can be reduced back to free cysteines.
Example: Insulin contains multiple disulfide bridges, as shown in the bovine insulin sequence.

Significance of Amino Acid Sequence

Sequence Determines Structure and Function

The unique sequence of amino acids in a protein determines its 3D structure, which in turn dictates its function. Knowledge of the sequence allows for:

Prediction of 3D structure
Understanding of catalytic mechanisms
Study of molecular pathology (disease-causing mutations)
Elucidation of evolutionary relationships

Example: Sanger's determination of the bovine insulin sequence was a landmark in understanding protein structure and function.

Peptide Bond Properties and Geometry

Planarity and Resonance of the Peptide Bond

The peptide bond exhibits partial double-bond character due to resonance, restricting rotation and resulting in a planar geometry involving six atoms (Cα, C, O, N, H, and the next Cα).

Resonance structures: Delocalization of electrons between the carbonyl oxygen and the amide nitrogen.
Planarity: The peptide bond is rigid and planar, with the six atoms lying in the same plane.
Trans vs. cis configuration: Most peptide bonds are in the trans configuration due to reduced steric hindrance between side chains.

Rotation Around Bonds: Phi (φ) and Psi (ψ) Angles

Although the peptide bond itself is rigid, rotation is possible around the N–Cα (phi, φ) and Cα–C (psi, ψ) bonds. The allowed values of these angles are limited by steric hindrance.

Ramachandran plot: A graphical representation of the permissible combinations of φ and ψ angles in a polypeptide chain.
Typical values: For α-helices, φ ≈ –60°, ψ ≈ –45°; for β-sheets, φ ≈ –120°, ψ ≈ +120°.

Secondary Structure: The α-Helix

Structure and Properties of the α-Helix

The α-helix is a common secondary structure in proteins, characterized by a right-handed coil stabilized by intra-chain hydrogen bonds.

Hydrogen bonding: The carbonyl oxygen of residue i forms a hydrogen bond with the amide hydrogen of residue i+4.
Dimensions: Each turn of the helix contains 3.6 residues and rises 0.56 nm (5.6 Å); each residue advances 0.15 nm (1.5 Å).
Dipole moment: The helix has a macrodipole, with a partial positive charge at the N-terminus and a partial negative charge at the C-terminus.
Amphipathic nature: Some α-helices have hydrophilic and hydrophobic faces, allowing interaction with both aqueous and lipid environments.
Disruptors: Proline (introduces kinks), isoleucine, threonine (branched at β-carbon), aspartic acid, asparagine (side-chain H-bond donors), and glycine (too flexible) can disrupt α-helix formation.

Secondary Structure: β-Strands and β-Sheets

Structure and Properties of β-Sheets

β-Strands are extended polypeptide chains that align side-by-side to form β-sheets, stabilized by inter-strand hydrogen bonds.

β-Strand: A nearly fully extended polypeptide segment; adjacent strands form a zig-zag or pleated pattern.
β-Sheet: Composed of two or more β-strands; can be parallel (strands run in the same direction) or antiparallel (strands run in opposite directions).
Hydrogen bonding: Occurs between the carbonyl oxygen of one strand and the amide hydrogen of an adjacent strand.
Distance: 3.5 Å between adjacent amino acids in a strand.
Side chain orientation: Alternates above and below the plane of the sheet.

Secondary Structure: Loops and Turns

Connecting Elements in Protein Structure

Loops and turns connect α-helices and β-strands, allowing the polypeptide chain to reverse direction and fold into compact structures.

Loops: Flexible regions, often found on protein surfaces, that can contain hydrophilic residues and vary in length (2–16 residues).
Turns: Short loops (≤5 residues), also called reverse turns or β-turns, stabilized by a hydrogen bond between the carbonyl oxygen of residue i and the amide hydrogen of residue i+3.
Function: Facilitate compact folding and are often involved in protein-protein or protein-ligand interactions.

Summary Table: Levels of Protein Structure

Level	Description	Stabilizing Interactions	Example
Primary	Linear sequence of amino acids	Covalent peptide bonds	Insulin sequence
Secondary	Local folding into α-helices and β-sheets	Hydrogen bonds	α-helix in myoglobin
Tertiary	3D folding of a single polypeptide	Hydrophobic interactions, disulfide bonds, ionic bonds, hydrogen bonds	Lysozyme structure
Quaternary	Association of multiple polypeptides	Non-covalent interactions, disulfide bonds	Hemoglobin tetramer