M121 Lec 5

Adaptive Immune Response - Antibody Structure and Function

Key Takeaways

• The interaction between antibodies and antigens is not covalent.

• Forces involved: Ionic bonds, Van der Waals forces, Hydrogen bonds, Hydrophobic interactions.

• These interactions contribute to antibody-antigen affinity.

Antibody Structure

• FAB region (Fragment Antigen Binding): Recognizes and binds to epitopes on antigens.

• FC region (Fragment Crystallizable): Binds to complement proteins and FC receptors on immune cells.

• Antibody Isotypes:

  • IgM (Mu) - Pentameric, low affinity but high valency.

  • IgE (Epsilon) - Involved in degranulation of mast cells and eosinophils.

  • IgG (Gamma) - Four subclasses, key for adaptive immunity.

  • IgA (Alpha) - Dimeric, joined by J-chain, essential for mucosal immunity.

  • IgD (Delta) - Role unclear but used as a marker for B cell maturation.

Effector Functions of Antibodies

1. Complement Activation: Classical pathway involving C1Q binding to FC region.

2. Opsonization: Enhancing phagocytosis by marking antigens.

3. Neutralization: Preventing pathogens from binding to host cells.

4. Degranulation: Triggering release of granules from mast cells, basophils, and eosinophils.

5. Antibody-Dependent Cellular Cytotoxicity (ADCC): Marking infected cells for killing by NK cells.

Antibody Valency and Epitope Recognition

• Antibodies recognize linear and discontinuous epitopes.

• Antigens can have multiple epitopes, leading to multiple antibodies binding.

• Higher valency improves immune response even if affinity is low.

• ELISA assays utilize this principle for protein quantification.

Antibody Effector Mechanisms

I. Complement Activation (Classical Pathway)

• Antibodies can activate the complement system, leading to various immune responses.

• Key Steps:

  1. Classical Pathway Activation:

     • IgM (most efficient) or two IgG antibodies bind to an antigen.

     • C1Q binds to the FC portion of antibodies.

     • C1R autoactivates and cleaves C1S, activating the cascade.

  2. Effector Mechanisms of Complement:

     • Recruitment & Inflammation: C5a and C3a act as chemoattractants for phagocytes.

     • Opsonization: C3b binds to pathogens, enhancing phagocytosis via CR1 receptors on macrophages.

     • Membrane Attack Complex (MAC): Forms pores in pathogen membranes, leading to lysis.

II. Opsonization

• Definition: The process of tagging pathogens for enhanced phagocytosis.

• Pathways for Opsonization:

  • Complement-Dependent: C3b binds to CR1 on macrophages to enhance phagocytosis.

  • Antibody-Dependent: Antibodies bind to pathogens, and their FC portion interacts with FC receptors on phagocytic cells.

    • FC Gamma Receptors (FCγRs) primarily bind IgG and enhance uptake by phagocytes.

• Functional Outcome:

  • Pathogens are ingested into phagosomes.

  • Phagosomes fuse with lysosomes (phagolysosome formation).

  • Degradation occurs via oxidative and non-oxidative mechanisms.

III. Neutralization

• Antibodies block critical functions of pathogens or toxins, preventing infection or damage.

• Mechanisms of Neutralization:

  1. Blocking Pathogen Adhesion:

     • Antibodies bind bacterial adhesins, preventing bacterial attachment to host cells.

  2. Toxin Neutralization:

     • Antibodies bind bacterial toxins, preventing them from interacting with host receptors.

  3. Blocking Viral Entry:

     • Antibodies bind viral surface proteins, preventing their interaction with host cell receptors.

IV. IgA and Mucosal Immunity

• Function of IgA in the Gut:

  • IgA is transported across epithelial cells via transcytosis.

  • Secreted IgA binds to mucin proteins in mucus.

  • Pathogens that bind IgA are swept away with mucus clearance.

• IgA in Viral Neutralization:

  • IgA binds to viral capsid proteins required for entry.

  • Prevents virus from infecting epithelial cells.

1. Neutralizing vs. Non-Neutralizing Antibodies

• Neutralizing antibodies prevent viral entry by blocking key receptors that viruses use to enter host cells.

• Non-neutralizing antibodies may still play a role in clearing the virus by other immune mechanisms but do not prevent initial infection.

• Example: If a person receives a vaccine and still gets infected, the vaccine likely did not generate strong neutralizing antibodies. However, it may have produced antibodies that aid in clearing the virus post-infection.

2. Degranulation and IgE-Mediated Responses

• Definition: Degranulation is the release of enzyme-filled granules by immune cells to combat pathogens.

• Key Cells Involved: Mast cells, basophils, eosinophils.

• IgE Function:

  • Unlike IgG, IgE is typically bound to the surface of mast cells via the FCε receptor.

  • When an antigen binds to IgE, it triggers cross-linking of FCε receptors, leading to mast cell degranulation.

  • Histamine Release: Causes strong contractions in tissue (e.g., sneezing, diarrhea) to expel pathogens, especially parasites.

  • Allergic Reactions: In developed regions where parasite infections are rare, IgE responses may be misdirected towards harmless antigens like pollen, leading to allergies.

3. Antibody-Dependent Cellular Cytotoxicity (ADCC)

• Definition: A mechanism where immune cells kill antibody-tagged infected or abnormal host cells.

• Key Players:

  1. Natural Killer (NK) Cells:

     • Recognize and bind to the FC portion of IgG via FCγRIII (CD16).

     • Release cytotoxic molecules (Perforin & Granzymes) to induce apoptosis in target cells.

  2. Macrophages: Can also mediate ADCC but are less dominant than NK cells.

• Steps in ADCC:

  1. Antibody binds to antigen on the infected or abnormal cell.

  2. NK cell binds to the antibody’s FC region via CD16.

  3. Cross-linking of CD16 triggers the NK cell to release perforin and granzymes.

  4. Perforin creates pores in the target cell membrane.

  5. Granzymes enter through these pores and trigger apoptosis (programmed cell death).

  6. The infected/cancerous cell is eliminated.

4. Immune Evasion & Vaccine Development

• Vaccine Goal: The most effective vaccines generate neutralizing antibodies to prevent infection rather than just aiding in virus clearance.

• Example Study Design:

  • Twins: One vaccinated, one not.

  • Vaccinated Twin: Produces neutralizing antibodies that bind to viral entry receptors, blocking infection.

  • Unvaccinated Twin: Lacks protective antibodies, allowing viral entry and severe infection.

• Cancer & ADCC:

  • Some cancer cells express embryonic proteins, making them look foreign to the immune system.

  • Monoclonal antibody therapies exploit ADCC by tagging cancer cells for NK cell-mediated destruction.

5. Allergy Treatments and Immunotherapy

• Oral Immunotherapy (OIT):

  • Gradual exposure to allergens to train the immune system to tolerate them.

  • Used to treat food allergies by slowly increasing antigen exposure.

• Desensitization Therapies:

  • Aim to shift immune responses from IgE-mediated hypersensitivity to tolerance.

  • Example: Microdosing allergens to suppress overactive IgE responses.

Summary

• Neutralizing antibodies are key for preventing infection, while non-neutralizing antibodies aid in viral clearance.

• IgE-mediated degranulation evolved for parasite defense but contributes to allergies in modern environments.

• ADCC is an essential immune mechanism for eliminating infected and cancerous cells.

• Vaccine efficacy depends on generating neutralizing antibodies.

• Immunotherapy can modulate the immune response to allergens and tumors.

Immunoglobulin (Antibody) Isotypes and Their Functions

1. IgM

   • First antibody produced in an immune response.

   • Major function: Complement activation via the classical pathway.

   • Requires at least two molecules for effective complement activation.

2. IgE

   • Binds to FC epsilon receptor on mast cells.

   • Major function: Degranulation, releasing histamine and other mediators.

   • Important in allergic reactions and parasite immunity.

3. IgG

   • Most abundant antibody in circulation.

   • Has four subclasses with multiple functions:

     • Complement activation (like IgM, requires at least two molecules).

     • Antibody-dependent cellular cytotoxicity (ADCC) via FC gamma receptor 3 (CD16) on NK cells.

     • Neutralization of pathogens by binding to viral or bacterial antigens and preventing entry into cells.

4. IgA

   • Specialized for mucosal immunity.

   • Undergoes gut transcytosis, allowing secretion into mucosal surfaces (e.g., gut, respiratory tract, mouth).

   • Present in secretions, such as breast milk, providing passive immunity to newborns.

5. IgD

   • Acts as a marker for B cell maturation.

   • Its exact effector functions in pathogen clearance remain unclear.

Antibody Structure and Antigen Binding

• Antibody structure consists of:

  • FC region (constant region) responsible for effector functions.

  • Fab region (antigen-binding site) responsible for antigen specificity.

• Variable regions

  • Composed of variable heavy (VH) and variable light (VL) chains.

  • Determines epitope recognition.

  • Sequence differences allow the immune system to recognize a vast array of pathogens.

• Complementarity Determining Regions (CDRs)

  • The specific binding surface that interacts with antigens.

  • Determines the complementarity between the antigen-binding site and the epitope.

  • Genetic rearrangement in these regions allows the immune system to generate millions of unique antibodies.

B Cell Maturation and Clonal Expansion

• B cell receptor (BCR) development

  • Once matured, each B cell produces only one unique set of heavy and light chains.

  • B cells clone themselves upon activation to generate identical daughter cells with the same receptor.

• Genetic Recombination in Antibody Generation

  • Exception to the central dogma (DNA → RNA → Protein).

  • Allows the immune system to generate millions of different antibodies despite having a limited number of genes.

  • Involves VDJ recombination, which introduces variability in the antigen-binding region.