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Home , Antibody, Idiotype, J chain

Chapter 4. Immunoglobulin Structure and Function

1 - Each heavy and light chain 2 is made up of a number of domains (= Ig folding or Ig domains ). * * - Light chains consist of 2 domains ( C and V ). - Heavy chains have 4-5 domains (depending on * the class of ) - Each domain is about 110 Heavy chain= 446 aa Light chain= 214aa amino acids in length and 2 1. Functional Regions 1 contains an intrachain bond between 2. Types of chains two cysteines about 60 3. Constant & amino acids apart. Variable regions 4.

-150,000 molecular weight Basic Antibody Structure • = cancerous plasma cells • Monomer = 150,000

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What is the difference?

- Constant (C) and Variable (V) regions

1 RECAP: - The Fc region plays NO role in binding. 100,000 MW 3 (Fab) 2 - breaks antigen molecules into 2 Fab fragments and an Fc fragment.

- Pepsin breaks antibody molecules into an F(ab’)2 fragment and a VERY SMALL pFc’ fragment. 1 Mercaptoethanol 2 - Mercaptoethanol treatment results in 2 heavy and 2 2 (50,0000) light chains 2 (25,000) 2 H + 2 L - Complexes of cross-linked by antigen are Papain 1 called “immune complexes”. 2 (45,000) 1 (50,000) 2 Fab + Fc

1. Constant region - amino Summary acid sequence in the C- terminal regions of the H and • Molecule consists of Constant and Variable regions 1 L chains is the same . for both Light and Heavy chains (C H, V H, C L, V L) 2 2. Variable region - amino • Ig molecule made of domains 1 3 acid sequence in the N- • Domains ~ 110 aa terminal regions of the H and • Each antigen-binding site is made up of the N- L chains is different . This terminal domain of the heavy and the light chains region provides antibodies with unique specificity. • IgM and IgE possess 4 C H domains (C H1-CH4) while IgG, IgA and IgD have 3 C H domains (C H1- 3. Hyper-variable regions are CH3). Hinge region is missing. regions within the variable • Hypervariable regions in the Variable regions of regions (greater specificities). both H and L chains.

Figure 3.3

-Within the variable domains are three regions of extreme Complementarity-Determining Regions , or CDRs. variability.

These are referred to as the hypervariable regions.

These regions of the variable domains actually contact the antigen.

They therefore make up the antigen-binding site.

These regions are also called the complementarity- Heavy Chain Light Chain determining regions , or CDRs.

2 H chain CDRs - A simulated antigen- RECAP: binding site showing - Antibodies are comprised of repeating 110 aa units referred to how the CDRs form as domains or Ig folds. points of contact with - The C-terminal domains are constant from antibody to the antigen. antibody (within a class).

- The constant region domains are responsible for all functions L chain CDRs of antibody other than antigen binding ( opsonization, ADCC, complement activation)  Biological Function!

- The N-terminal domains are variable from antibody to antibody and are referred to as “variable domains ”.

- The variable domains contain 3 hypervariable regions - the CDRs .

- The CDRs of the V domains in both H and L chains make up the antigen-binding site .

Fc γ receptors enhance of foreign cells/particles Antibody-Mediated Effector coated with IgG

Functions Antibody made in response to foreign cells (cells/viral • Binding to Antigen particles/ etc) will bind to those cells. • OPSONIZATION: FcR in and Macrophages (and ) possess receptors for the Fc neutrophils region of IgG. • COMPLEMENT ACTIVATION: IgG and IgM Binding of Fc receptors to antibody bound to cells/particles facilitates and increases phagocytosis of • ADCC – NK cells trough FcR cells/particles. • CROSSING EPITHELIAL LAYERS – IgA (but also IgM) • CROSSING - IgG

ADCC - Antibody-dependent cellular - mediated by IgG

Antibody made in response to foreign cells (cells/viral particles/bacteria etc) will bind to those cells.

Cells of the innate (neutrophils, , macrophages, NK cells) possess receptors for the Fc region of IgG.

These cells bind to antibody on the surface of foreign cells Monomer, and release lytic compounds Dimer,  . and Pentamer

Kuby Figure 14-12

3 Structural Variants of the Basic Immunoglobulin Molecule Relative abundance in normal : Different heavy chains can be used There are five major types of heavy chain --> five major classes IgG 8 - 16 mg/ml (isotypes ) of antibody IgA 1.4 - 4 mg/ml

- gamma --> IgG (in humans 4 subclasses: IgG1, IgG2, IgG3, IgG4) IgM 0.5 - 2 mg/ml - mu --> IgM - alpha --> IgA (in humans, 2 subclasses: IgA1, IgA2) IgD 0.003 - 0.04 mg/ml - delta --> IgD - epsilon --> IgE IgE 17 - 450 ng/ml (<0.0005 mg/ml) IgD The function of antibody varies depending on which heavy chain is IgM used. IgE IgA IgG IgA IgM IgD IgE

IgG IgG IgM IgA IgD IgE

-Most abundant in secondary responses -Crosses placenta (FcRn) -Complement activation -Binds to FcR in

Figure 3.15a

Crosses placenta Crosses placenta Crosses placenta Complement Activator Complement Activator Complement Activator Fc binding Fc binding - Best Complement activation - First Ab produced in neonate - First antibody produced after challenge - Mucosal transport (to some degree) - Monomer on B cells - : polymeric

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- Dimer in mucosal - Mucosal transport Secretory Component - Monomer in circulation - J chain (polymeric) and Secretory components

Role of IgE in allergic reactions IgD IgE antibodies mediate the immediate- (allergic) reactions that - Role unknown are responsible for symptoms of hay fever, , and anaphylactic - Present on the shock. surface of MATURE IgE binds to Fc receptors on the membranes of and B cells  Marker!! tissue mast cells .

Cross-linkage of receptor-bound IgE molecules by antigen () induces of basophils and mast cells.

A variety of pharmacologically active mediators present in the granules are released, giving rise to allergic manifestations

SUMMARY Antigenic Determinants on - IgA and IgM are secreted across epithelial surfaces Immunoglobulins - IgG, IgD and IgE can be found only within the • Abs are and themselves very body - in serum or lymph . immunogenic - IgA and IgM are also found in serum and lymph BUT • on immunoglobulins are divided IN ADDITION can also be found in secretions such as mucous secretions, saliva and . into: – ISOTYPIC - The IgA and IgM found in external secretions differs from that found in serum by the presence of an – ALLOTYPIC additional component referred to as the "secretory – IDIOTYPIC component".

- This component is acquired as the IgA or IgM is transported across the epithelial barrier.

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Constant region determinants that define each antibody Allelic variation (Allotypes): IgG of a particular class may be class and subclass slightly different between individuals ( e.g. variation in the IgG sequence) The function of antibody varies depending on which heavy chain Note: This type of variation has no effect on antibody is used. function.

RECAP - Sequence variation in antibodies:

1. Different light changes - no significant functional effect

2. Different heavy chains - very significant functional effect - isotypic variation

3. Allelic variation between individuals - no large functional effect - allotypic variation Generated by variation in amino acid sequence in the V H and VL. Most exactly, in the CDRs in the V regions 4. Variation in the antigen-binding site - idiotypic variation Variation in the antigen binding site ( )

Remember: = Ag binding site

B Cell Receptor (BCR): Ig Superfamily - Short cytoplasmic tail (3- 28 aa) ….signaling? • Divergence from a common ancestor coding - Signaling through a for 110 aa. homodimer, Ig-α and Ig-β • A member MUST have a “typical” Ig domain or fold  110 aa with an intra chain disulfide bond - Ig molecule + Ig-α/Ig-β is 50-70 aa apart. the BCR • Most members do not bind Ag!! Then, they must - The homodimer molecule facilitate interaction with surface is member of the Ig • You must know members with roles in: a) immune superfamily group function, b) Receptor/, and c) Adhesion

6 Receptors Immune Function

Neonatal

Monoclonal Antibodies

• Kohler & Milstein 1975 • Fusion of normal, activated and (cancerous ) • Hybrid: immortal, secrete Ab, hypoxanthine

Plasmacytoma VS B cell RESULTS: • Plasmacytoma: – Cancerous plasma cell (Immortal) – Does not secrete Abs B cell Hybrid Plasmacytoma – Lacks HGPRT ** • Normal spleen B cell Die in culture Immortal, Secretes Lacks HGPRT – Limited life span Ab, Possess – Secretes Abs hypoxanthine (HGPRT) – Possess HGPRT

7 1 Applications? 2 3 4 • Diagnosis • Research 5 • Treatment • Affinity VS Affinity (polyclonal Ab ) = high because of multiple epitopes Avidity (monoclonal Ab ) = low affinity but high avidity because of strong - Ab interaction

IgG - Most abundant Ig of internal body fluids (serum, extracellular fluids) - combats and within the body tissues. The End IgA - Most abundant Ig in mucous secretions - protects external surfaces of the body IgM - The first class of antibody produced during an . Present both in internal body fluids and in secretions.

IgD - Functions not well defined. Found mostly on the B cell plasma membrane

IgE - Increases during parasitic . Causes symptoms of .

IgG IgA IgM IgD IgE

Complement fixation by ++ - +++ - - classical pathway

Ability to cross the placenta ++ - - - - Binds to mast cells and - - - - +++ basophils Binds to macrophages +++ + - - + and polymorphs

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