PHARM 310: LEC 2 - Antibody Structure and Function

Antibody Structure

• Antibodies are Y-shaped glycoproteins made of 4 polypeptide chains: 2 heavy (50 kDa each) and 2 light (25 kDa each) chains.
• Chains are connected by disulfide bonds, consisting of a variable region (N-terminus) and constant regions.

Antibody Specificity and B-cell Activation

• Each antibody (Aby) has a unique specificity; the human antibody repertoire contains approximately 10^16 variations.
• B-cells express a single immunoglobulin (Ig) type on their surface and differentiate into plasma cells upon antigen (Ag) binding.

Antibody Functional Regions

• Protease digestion reveals antibody functional regions:
  • Papain cleaves antibodies into 2 Fab (Fragment antigen binding) and 1 Fc (Fragment crystallizable).
  • Pepsin digests the Fc fragment, yielding F(ab')2.
• The Fc region is crucial for opsonization.

Antibody Isotypes

• Immunoglobulin isotypes (classes) differ in the constant regions of heavy and light chains; five major isotypes exist: IgG, IgA, IgM, IgE, IgD (GAMED).
• Heavy chains are denoted by Greek letters: γ, α, μ, ε, δ; light chains have two types: Kappa (κ) and Lambda (λ).

Variable and Constant Domains

• Immunoglobulin domains range from 100-110 amino acids.
• The variable region (V-region) includes VH (heavy chain) and VL (light chain) domains, while the constant regions have limited sequence diversity.

Antigen-binding Site

• Known as the paratope, consists of framework regions (FR) and hypervariable (HV) loops called complementarity determining regions (CDRs).
• CDRs are responsible for the specificity of antigen binding.

Antigenic Determinants (Epitopes)

• Antibodies bind antigens at specific sites known as epitopes, often composed of carbohydrates, proteins, or both.
• Most antigens present multiple epitopes, making them multivalent.

Binding Mechanism

• Antibody-antigen interactions exhibit molecular complementarity through non-covalent binding (ionic, hydrogen bonds, hydrophobic effects, van der Waals attractions).
• The "Lock and Key" model indicates specific interactions based on shape and charge compatibility.

Gene Structure and Protein Coding

• Antibodies are proteins coded by segments of DNA (genes); base pairs include A (Adenine), T (Thymine), C (Cytosine), G (Guanine).
• Eukaryotic DNA contains coding (exons) and non-coding (introns) segments.

Transcription and Translation

• Primary RNA transcripts from antibodies include introns, while the processed mRNA (messenger RNA) excludes them, coding for polypeptides.

Post-translational Modifications

• Polypeptide glycosylation involves sugar attachment in the Endoplasmic Reticulum and Golgi, specifically N-glycosylation and O-glycosylation.
• Glycosylation is critical for protein folding and localization.

Immunoglobulin Diversity

• Antibody diversity is generated prior to antigen encounter via gene segmentation and recombination.
• Ig genes are fragmented in all cells except B-cells, giving rise to a unique germline configuration.

Gene Segment Recombination

• RAG-1 and RAG-2 enzymes mediate gene segment recombination in B and T-cells.
• Ig constant regions are prepared for transcription, while variable regions require RAG-mediated rearrangements.

Germline Ig Genes and Diversity

• Ig genes undergo rearrangement to produce multiple combinations of V, D, and J segments, significantly enhancing diversity.

Affinity Maturation and Isotype Switching

• Somatic hypermutation in B-cells improves antibody specificity over time.
• Isotype switching allows B-cells to produce different antibody isotypes (IgM, IgG, IgA, IgE) through DNA recombination, altering the immune response without changing the V-region specificity.

Isotype Switching Mechanism

• IgM and IgD are initially expressed on B-cells via differential mRNA splicing.
• Subsequent switch to IgG, IgA, or IgE allows diversification of immune function based on the need.

Summary of Antibody Attributes

• Each antibody class features distinct functions facilitated by different constant regions, while the V-region specificity remains consistent across isotypes.

This quiz explores the structure and function of antibodies, including their varying isotypes and specificity. Understand how antibodies are structured from polypeptide chains and their roles in B-cell activation and immune response. Delve into the functional regions of antibodies and their significance in opsonization.