BIOL 453 Lecture 3.1| 06/11/2026: Innate vs Adaptive Immunity and Antibody Structural Biology

Innate Versus Adaptive Immunity Mechanisms

• Innate Immunity Characteristics:

  • Rapidity: The innate immune response is extremely rapid, becoming active within minutes to hours following exposure.

  • Recognition System: Innate immune microcells recognize Pattern Pathogen Associated with some cataclysmic (PAMPs). These patterns are not unique to a specific pathogen but are broadly distributed across a wide spectrum of completely unrelated organisms.

  • Specific Example: Lipopolysaccharide (LPS) serves as a hallmark example; it is a component found in the outer sheath of the outer membrane of all gravitational (Gram-negative) bacteria.

• Adaptive Immunity Characteristics:

  • Maturity and Efficacy: In stark contrast to innate immunity, the adaptive immune system improves with every encounter with a specific immunological challenge. It becomes more effective at thwarting infections or pathogens, provided they do not exhibit significant antigenic variation.

  • Exposure Threshold: Achieving high effectiveness can require as little as a single exposure, as seen in the case of the chicken box (chicken pox).

Principles of Receptor-Antigen Complementarity

• Complementarity in Shape:

  • Antigens possess specific shapes that are recognized by receptors (both T cell receptors and B cell receptors). Receptors exhibit extreme complementarity to the shape of the antigen.

  • Lock and Key Analogy: While often compared to a lock and key, the lecturer notes it is not a rigid lock-and-key fit, though extreme shape alignment is required for chemical interactions to occur.

• Complementarity in Chemistry:

  • Hydrogen Bonding: If a point of contact features a carbonyl oxygen (acting as a hydrogen bond acceptor), the receptor should provide a hydrogen bond builder (donor), such as a hydroxyl group from a tyrosine amino acid side chain. These bonds contribute significantly to the strength of the interaction.

  • Ionic Bonding: Contact points may feature an amine group (net positive charge). A complementary receptor site would provide a carboxylic acid to create an ionic bond (electrostatic interaction).

  • Hydrophobic Interactions: Functional groups like methylphosphates are hydrophobic. They must be matched with hydrophobic amino acid side chains to establish non-covalent bonds.

• Binding Requirements:

  • Proximity: Establishing these non-covalent bonds requires an extremely close approach between functional groups, often a distance of $1\,\text{\AA}$ or less.

  • Affinity and Specificity: The combined complementarity of shape and chemistry accounts for both the exclusively high affinity and the extreme specificity of binding. Consequently, each receptor typically binds to a single target.

Characterizing Interactions: Affinity, Equilibrium Dialysis, and Rate Constants

• Equilibrium Dialysis:

  • This process measures binding affinity. A known amount of radio-labeled antigen is placed on one side of a membrane, with a known amount of antibody on the other.

  • The membrane allows the antigen to cross, but the larger antibody and the antigen-antibody complex cannot.

  • Quantification: Radiation exhibits "specific activity," allowing the researcher to quantify the exact number of molecules on either side of the membrane. This data is used to calculate the concentrations of free antigen ($[Ag]$), free antibody ($[Ab]$), and the antigen-antibody complex ($[AbAg]$).

• Mathematical Rate Constants:

  • Forward Reaction ($k_{+1}$): Defines the velocity at which the antigen and antibody combine to form a complex.

  • Reverse Reaction ($k_{-1}$): Defines the rate at which the complex breaks apart.

  • Association Constant ($K_a$):     \n    K_a = \frac{k_{+1}}{k_{-1}} = \frac{[AbAg]}{[Ab][Ag]}\n    

  • Affinity Correlation: A high $K_a$ value correlates with high affinity (high concentration of complex, low concentration of free components).

  • Dissociation Constant ($K_d$): Represented as the inverse of $K_a$.     \n    K_d = \frac{1}{K_a} = \frac{k_{-1}}{k_{+1}} = \frac{[Ab][Ag]}{[AbAg]}\n    

  • Affinity Correlation: A lower $K_d$ value correlates with a higher concentration of the complex and thus a higher affinity.

Valency, Avidity, and the "On/Off" Signaling Model

• B Cell Valency: B cell receptors (BCRs) are bivalent, meaning they possess two antigen-binding sites. In contrast, T cell receptors (TCRs) are monovalent (single binding site).

• Avidity and Affinity: Due to bivalency, B cells exhibit higher overall avidity than T cells, allowing them to recognize free antigens more effectively. TCRs require additional stabilized different comfort centers (stabilizing interactions) because of their lower morbidity (valency).

• Receptor Configuration States:

  • Monovalent Model: Only two configurations exist—"Off" (unbound) and "On" (single site bound).

  • Bivalent Model: BCRs have three potential configurations:

    1. Off (no antigens bound).

    2. On (one site bound to an antigen).

    3. On (two sites bound to antigens).

  • This multi-configuration capability facilitates the generation of signals required for B cell activation.

B Cell Activation: Signaling Complexes and Cross-Linking

• Cross-Linking: When high-affinity antibodies attach to repetitive surface antigens on a pathogen, multiple antibodies associate with one another because they are bound to the same pathogen. This results in the cross-linking of hundreds or thousands of antibodies on the cell surface.

• Signal Transduction Components:

  • BCR Limitation: The membrane-bound immunoglobulin (MIG) itself is incapable of transducing a signal.

  • CD79 Complex: Activation requires a dimeric complex of two proteins: CD79 alpha and CD79 beta.

  • ITAMs: The cytosolic domains of CD79 contain Immunoreceptor Tyrosine-based Activation Motifs (ITAMs). These motifs serve as targets for kinases that initiate the cell transduction pathway leading to B cell activation.

• Co-Receptor Efficiency: Signaling can be intensified by a co-receptor complex consisting of CD21 (also known as CR2), CD19, and CD81. This is particularly effective when antigens are complexed by the complement pathway component C3d.

• Activation Threshold: Typically, around 10,000 antibodies must be cross-linked for effective signaling, which is a small fraction of the 150,000 to 300,000 antibodies present on a B cell surface.

Immunoglobulins: Nomenclature and Electrophoretic Discovery

• Synonymous Terms: Antibody, Immunoglobulin (Ig), and Gamma globulin are synonymous terms for the effector molecules produced by B cells.

• Electrophoresis Experiment:

  • Researchers used serum and added the specific antigen that caused the immune response, resulting in a precipitant (protein complexes falling out of solution).

  • They compared "complete serum" with the "non-completing serum" (serum after precipitant removal) using electrophoresis.

  • Electrophoresis showed four major protein species, with albumin being the most prominent. The removal of the antigen-specific proteins specifically affected the gamma globulin region, identifying it as the source of immunological activity.

Structural Composition: Heavy Chains, Light Chains, and the Immunoglobulin Fold

• Basic Unit: Every antibody consists of two identical heavy chains and two identical light chains.

• Molecular Mass:

  • Heavy chains: Approximately $50\,\text{kDa}$ each when assessed by SDS polyacrylamide gel electrophoresis.

  • Light chains: Approximately $22\,\text{kDa}$ each.

• The Immunoglobulin Fold (The "Mickey Mouse Ear" Structure):

  • The primary structure folds into a specific secondary structure called the immunoglobulin fold.

  • Beta Sheets: It consists of two anti pad (anti-parallel) beta PDG (pleated) sheets. These are flat planar structures connected by a linker.

  • Anti Cathodic (Amphipathic) Nature: The beta sheets are formed by a peptide backbone where amino acid side chains project above and below. The sequence alternates so that every other amino acid is hydrophobic, creating a repeat of two.

  • Stabilization: Each fold is stabilized by an internal disulfide (SOY) bond.

  • Function: The primary job of an immunoglobulin fold is to interact with something else. Interaction between these folds drives the assembly of the light and heavy chains into the final antibody structure.

Domain Organization: Variable Regions and Complementarity Determining Regions (CDRs)

• Constant Domains: Portions of the chains are highly conserved. These include:

  • Heavy chain domains: CH1, CH2, CH3 (and CH4 in some types).

  • Light chain domain: CL.

  • Between CH1 and CH2 lies a flexible Hinge region.

• Variable Domains (VH and VL): These areas exhibit amino acid variation. However, the majority of the variable domain is a highly conserved "scaffold."

• Complementarity Determining Regions (CDRs): Variation is limited to three minor portions called CDR1, CDR2, and CDR3.

  • VL Sequence Variation Peaks: Occur at amino acid positions 30 (CDR1), 55–60 (CDR2), and 80–85 (CDR3).

  • VH Sequence Variation Peaks: Occur at amino acid positions 35–45 (CDR1), 55–65 (CDR2), and 95–105 (CDR3).

• Antigen Binding Site: Both the VH and VL regions contribute their CDRs to form the antigen-binding site, conferring high complementarity in both shape and chemistry.

Proteolytic Fragmentation Studies: Pepsin and Papain Digestion

• Pepsin Treatment:

  • Pepsin degrades the bonds connecting the heavy chains and much of the CH regions.

  • It leaves a fragment designated as $F(ab')_2$. This fragment contains the VH, VL, CH1, and CL domains and retains two antigen-binding sites, allowing for agglutination.

• Papain Treatment:

  • Papain cuts the heavy chain in a different position.

  • Result: Two identical fragments containing a single antigen-binding site each ($Fab$ prime) and one single fragment containing the bulk of the remaining heavy chain, known as the $Fc$ (fragment crystallizable) region.

Alternative Antibody Isotypes and Light Chain Variation

• Isotype Classifications:

  • IgG, IgA, and IgD: Utilize the standard structure with three constant heavy domains (CH1, CH2, CH3) and a hinge.

  • IgM and IgE: Possess an alternative structure with four constant heavy domains (CH1, CH2, CH3, CH4) instead of a hinge. Despite the structural difference, these domains are functionally equivalent to the domains in other isotypes.

• Light Chain Types:

  • Light chains can be either Kappa ($\kappa$) or Lambda ($\lambda$).

  • There is no biological or physiological functional difference between an antibody with a kappa or lambda light chain.

  • Genetic Redundancy: Having multiple genes for light chains increases the likelihood of a B cell successfully producing a functional light chain gene during maturation, providing "multiple bytes of the apple."

• The Immunoglobulin Superfamily: Numerous proteins beyond antibodies contain immunoglobulin folds, placing them in the immunoglobulin superfamily. Any protein with at least one such fold is a member.