ANTIBODY STRUCTURE AND FUNCTION TODAY ANTIBODY STRUCTURE AND FUNCTION

The BCR directly recongnizes its cognate antigen Antigens are detected by specialized T cell receptors (TCR) or B cell receptors (BCR). APC T cells require antigen presentation by dedicated antigen presenting cells (APC) The BCR directly recongnizes its cognate antigen B cell T cell An antigen is anything that activates a lymphocyte. It can be a peptide, recognized by T cell receptor on a T cell, or a large three dimensional object, recognized by a B cell receptor on a B Cell.

LYMPHOCYTE ACTIVATION IS TRIGGERED BY ANTIGEN RECOGNITION MICROBE B CELL Antigen recognition is mediated by structurally related cell surface molecules: membrane-bound antibodies on B lymphocytes and T cell receptors on T lymphocytes. Antigen receptors distinguish many, and often chemically related, structures. Antigen receptors are clonally distributed: Each clone has a unique receptor distinct from all other receptors Clone: A parental lymphocyte and all its progeny.

BCR/TCR ACTIVATION 1. Antigen receptors are non-covalently associated with cellular signaling proteins in the BCR or TCR complex. BCR COMPLEX Extracellular BCR 2. When adjacent antigen receptors bind two or more antigens the receptors are pulled together into an aggregate. Intracellular 3. Cross-linking brings signaling proteins together and initiates signal transduction. 4. As the signaling proteins are the same in each clone, the transduced signal is also the same.

B CELLS 1. B CELLS ARISE FROM BONE MARROW STEM CELLS. 3. B CELLS RECIRCULATE THROUGH LYMPHOID ORGANS IN SEARCH OF ANTIGEN. 2. MATURE B CELLS EXIT BONE MARROW AND ENTER THE CIRCULATORY SYSTEM. 1. B CELLS ARISE FROM BONE MARROW STEM CELLS.

B CELLS 1. BCR detect three-dimensional antigens. 2. B cell activation does not require antigen presentation. 3. B cells are active only against extracellular antigens.

B CELLS AND ANTIBODIES Facts about the BCR and its relationship to secreted immunoglobulin (Ig) 1. Each B cell makes One specificity of antibody (Immunoglobulin (Ig)). The antigen receptor is membrane bound on a naïve B cell. 2. Ag plus second signal stimulate B cell. 3. Active B cell secretes antibody into the blood and lymph or patrolling mucosal surfaces.

B CELLS AND ANTIBODIES Naive Active BCR Secreted antibodies In the transition from naïve to active B cells, the B cells secrete a modified version of the B cell receptor. This molecule is known as an antibody. An antibody is a secreted version of a BCR with identical antigen specificity. This means that the antibody targets the exact same antigen that initially stimulated the BCR. The BCR and corresponding antibody share IDENTICAL antigen specificities.

What are Antibodies? Antibodies (A.K.A. Immunoglobulin) are a type of glycoprotein produced by B lymphocytes. Antibodies bind antigen with a high degree of specificity and affinity. Antibodies recognize a variety of three-dimensional shapes (amino acids, lipids, carbohydrates, etc).

ANTIBODY STRUCTURE LIGHT CHAIN HEAVY CHAIN An antibody consists of four polypeptides: two identical light chains and two identical heavy chains form a Y-shaped molecule. Each light chain is connected to a heavy chain by a disulfide bond and the two heavy chains are connected by two disulfide bonds. Antigen binding and effector domains are separated by a hinge region. The hinge region allows the two antigen binding domains to move, enabling them to bind antigens that are separated by varying distances.

ANTIBODY STRUCTURE ANTIGEN RECEPTORS HAVE DISTINCT FUNCTIONAL DOMAINS 1. The variable (V) region varies between clones and is involved in antigen recognition. 2. The constant (C) region is conserved among clones and is required for structural integrity and effector functions. Every antibody (secreted) is composed of the variable region and the contant region, a heavy chain (which contains both) and a light chain (which contains a variable region).

ANTIBODY STRUCTURE The unique structural architecture of antibodies allows multiple, highly diverse antigens to induce identical effector functions. Antigen Variable region Constant region IMMUNITY Different variable regions bind different antigens. Identical constant regions induce identical responses.

ANTIBODY STRUCTURE Each Heavy chain contains one V domain and at least three C domains. Each Light chain contains one V domain and one C domain. Each domain folds into a characteristic 3-D shape: The Immunoglobulin (Ig) domain. VL VH Antibodies bind antigens through the variable regions of the light and heavy chains.

BINDING OF ANTIGEN BY ANTIBODY - TERMS AND CONCEPTS Epitope: The parts of an antigen recognized by an antibody are called epitopes. Epitopes can be recognized on the basis of sequence or shape. Epitope Antibodies bind antigens by reversible non-covalent interactions. Affinity: The strength with which one antigen-binding surface of an antibody binds an antigen. Avidity: Each Isotype has between 2 and 10 antigen binding sites. Therefore, each antibody can bind 2 to 10 epitopes of an antigen, as long as identical epitopes are sufficiently close together, e.g. microbial cell surface proteins. In this case the binding is much greater than the affinity of a single antibody-antigen bond and is called the avidity.

Specific Antibody (Ig) Molecules Are Tailored to Exactly Fit A 3 D Surface Site On An Antigen. Epitope It looks as if the antibody was made specifically (custom made) for the antigen. Influenza coat protein Ig

FEATURES OF ANITBODY-MEDIATED 1. Antibodies recognize a large array of 3-D structures. 2. Each clone is specific for a single antigen. 3. Antigen recognition is mediated by specific domains of the antibody. 4. Signaling triggers B lymphocyte activation. FEATURES OF ANITBODY-MEDIATED ANTIGEN RECOGNITION

B CELL ACTIVATION B cell activation requires two signals Active B cell 1. Antigen 2. PAMP PRR BCR 1. Antigen A. T Cell-independent. 2. PAMP BCR PRR B cell 1st signal: BCR detects Ag. 2nd signal: PRR detects PAMP. CD4+ T cell B. T Cell-dependent. Cytokines 1. Ag The requirement for a second signal is the equivalent of a safety mechanism. Lymphocytes are only activated upon independent confirmation of an ongoing infection. This safety considerably lowers the potentially lethal risk of auto-immune responses. 1st signal: BCR detects Ag. BCR B cell B cell internalizes Ag and presents to CD4+ T cell (B cell as APC). MHC II/Ag 2nd signal: T cell produces cytokines that activate B cell.

IMMUNOGLOBULIN ISOTYPES There are five types of heavy chain that differ in their C region. Each class is an Isotype. Each Isotype differs in their physical/biological properties and effector functions. IgA. Two IgA molecules joined by J chain. The J chain facilitates transport of IgA across mucosal epithelia. The J chain also facilitates transfer of IgA to newborns to confer neonatal passive immunity. The J chain is an immunoglobulin part (only one type) which allows the chain to be transported across a mucosal epithelium.

IMMUNOGLOBULIN ISOTYPES IgD. Function unknown. Restricted to membrane and not expressed on active B lymphocytes. IgD knock-out mice do not have any apparent defects. The function of IgD is poorly understood, as deletion of IgD does not have any obvious defects.

IMMUNOGLOBULIN ISOTYPES Secreted as a monomer. Antigen IgE MAST CELL Fc Receptor IgE binds the Fc receptors of mast cells. IgE is involved in mast cell activation. IgE binds mast cells (and eosinophiles) where it waits for antigen. This is different from IgG, which must bind antigen before cells. When an antigen cross-links IgE, it causes mast cell degranulation. When IgE is cross-linked by antigen, it triggers mast cell degranulation - allergic response.

IMMUNOGLOBULIN ISOTYPES Secreted as monomer IgG can bind and neutralize toxins. IgA and IgG can bind and neutralize toxins.

IMMUNOGLOBULIN ISOTYPES Opsonization: IgG coats pathogens and prevents them from entering host cells. Infectious pathogen entering host cell to propagate IgG can coat (“opsonize”) microbes and thereby interfere with microbial infection of host cells. Pathogen cannot enter cell when coated (“opsonized”) with antibody

IMMUNOGLOBULIN ISOTYPES IgG Ag Phagolysosme FcR Macrophage MHC II/peptide Opsonized microbes are targets for phagocytosis by macrophages. This slide provides an excellent example of interplay between innate and adaptive arms of the immune system. Antibodies (adaptive) opsonize a microbe and thereby target it fro phagocytosis by a macrophage (innate). Antigens from the phagocytosed microbe are then presented on MHC II molecules to CD4 T cells (adaptive). The CD4T cells in turn can activate additional B cells (adaptive) that present the correct antigens or macrophages (innate) that present the correct antigens. Note how only B cells and T cells that respond to antigens from the infecting microbe are activated. This limits the response to the infection microbe. 1. IgG-Ag binds the Fc receptor of macrophages. 2. Ag is internalized through phagocytosis. 3. Microbe is degraded in the phagolysosome. 4. Microbial peptides are presented on Class II MHC molecules - CD4 T cell activation!

IMMUNOGLOBULIN ISOTYPES Fc receptors also mediate Antibody-Dependent Cellular Cytotoxicity (ADCC) Natural Killer Cells Ig Target Ag 1. Ig binds Ag on surface of target cell. Another case of adaptive activating innate! Here, the target cell is opsonized with antibodies (adaptive) and thereby targeted for destruction by NK cells (innate). 2. Fc Receptors on Natural Killer cells bind Fc of Ig. 3. Cross-linking of Fc receptors signals to the NK cell to kill the target. 4. Target cell dies by apoptosis

IMMUNOGLOBULIN ISOTYPES 10 different Ag binding sites - very high avidity! IgM exists as pentamers As with IgA, it has a J chain for secretion. IgM is the first antibody expressed in mature B cells. IgM is the first antibody produced by naïve B cells. It has very high avidity (10 antigen binding sites) and is involved in complement activation. Important for activation of the complement pathway. Appears early after infection and usually not seen upon re-infection.