Chapter 4 antibody structure and function JOANNAISMYNAME

IMMUNOGLOBULINS are glycoproteins found in the serum portion of the blood Composed of 82% - 96% polypeptide and 2% - 14% carbohydrate Humoral branch of the immune response Primary role is antigen recognition and in biological activities related to immune response and complement activation Has five major classes (IgG, IgM, IgA, IgD, IgE)

IMMUNOGLOBULIN: TERAPEPTIDE STRUCTURE Basic structure is a tetrapeptide: two H and L chins linked by disulfide bonds Intrachain disulfide bonds create flooded regions or domains The amino-terminal end of each chain is a variable region, while the carboxy-terminal end is one or more constant regions Pepsin digestion yields 𝐹(𝑎𝑏) 2 fragment, with all the antibody activity, and an 𝐹𝐶 1 fragment Papain digestion yields two 𝐹 𝑎𝑏 fragments and an 𝐹𝑐 portion

IMMUNOGLOBULIN: TERAPEPTIDE STRUCTURE

Papain cleavage

Pepsin digestion

Nature of light chains Bence-Jones Proteins Kappa (Є) Lambda (λ) Constant region is the C-terminal end and contains similar amino acids for each class of antibody. Variable region includes 110-130 amino acids of the light and heavy chains, and is responsible for binding to antigen. This part of the antibody shows variations in amino acids when the specificity of the antibody for antigen is changed.

Heavy chain sequencing Variable domain – first 110 amino acids at the amino-terminal domain The remaining amino acid can be divided up into three or more constant regions with very similar sequences, 𝐶 𝐻 1, 𝐶 𝐻 2, 𝐶 𝐻 3. IgG - Gamma (γ) heavy chains IgM - Mu (μ) heavy chains IgA – Alpha (α) heavy chains IgD – Delta (δ )  heavy chains IgE – Epsilon (ε ) heavy chains

ANTIGENIC PROPERTIES OF ANTIBODIES Isotype Unique amino acid sequence that is common to all immunoglobulin molecules of a given class in a species Allotype Found in the constant region and are inherited in simple Mendelian fashion Occur four in IgG subclasses, in one IgA subclss and in the kappa light chain Idiotype Found in the amino-terminal ends of both L and H chains Essential to the formation of the antigen-binding site Together they serve as antigen-recognition unit

Antigenic Properties of antibodies

Hinge region flexible amino acid stretch in the central part of the heavy chains of the IgG and IgA immunoglobulin classes, which links these 2 chains by disulfide bonds High content of proline and hydrophobic residues Flexibility assists inititiation of the complement cascade rich in cysteine and proline amino acids, extremely variable in amino acid sequence, no resemblance to any other immunoglobulin region Gamma, delta and alpha chains have hinge region Mu and epsilon chains do not have hinge region

Carbohydrate functions All types of immunoglobulins contain a carbohydrate portion located between 𝐶 𝐻 2 domains of the two H chains They function as: 1. Increases the solubility of the immunoglobulin 2. Provides protection against degradation 3. enhances functional activity of the 𝐹 𝐶 domains

3D antibody structure

3D antibody structure Basic four-chain structure is folded into compact globular subunits Intrachain of disulfide bonds stabilize globular regions Beta-pleated sheet Within each regions or domains, polypeptide chain is folded back and forth on itself Immunoglobulin Fold Cylindrical structure produced when folded domains of the H chains line up with those of the L chains

3D antibody structure Hypervariable regions Strategic locations wherein antigen binds to a small number of amino acid and then captured within the barrel Comparisons of the amino acid sequences of the variable regions of immunoglobulins show that most of the variability resides in three regions called the hypervariable regions Complementarity-determining regions (CDRs) found in both the H and the L chains Occur as loops in the folds of ariable regions Antigen binds in the middle of CDRs

Types of imMunoglobulin IgG Structure:  All IgG's are monomers (7S immunoglobulin). The subclasses differ in the number of disulfide bonds and length of the hinge region. Properties most versatile immunoglobulin because it is capable of carrying out all of the functions of immunoglobulin molecules. a) IgG is the major Ig in serum - 75% of serum Ig is IgG b) IgG is the major Ig in extra vascular spaces

Subclasses a) IgG1 - Gamma 1 heavy chains 67% b) IgG2 - Gamma 2 heavy chains 22% c) IgG3 - Gamma 3 heavy chains 7% Largest hinge region d) IgG4 - Gamma 4 heavy chains 4%

Types of imMunoglobulin c) Placental transfer - IgG is the only class of Ig that crosses the placenta. Transfer is mediated by a receptor on placental cells for the Fc region of IgG. Not all subclasses cross equally well; IgG2 does not cross well. d) Fixes complement - Not all subclasses fix equally well; IgG4 does not fix complement e) Binding to cells - Macrophages, monocytes, PMNs and some lymphocytes have Fc receptors for the Fc region of IgG. Not all subclasses bind equally well; IgG2 and IgG4 do not bind to Fc receptors. A consequence of binding to the Fc receptors on PMNs, monocytes and macrophages is that the cell can now internalize the antigen better. The antibody has prepared the antigen for eating by the phagocytic cells. The term opsonin is used to describe substances that enhance phagocytosis. IgG is a good opsonin. Binding of IgG to Fc receptors on other types of cells results in the activation of other functions.

Functions of immunoglobulin g Providing immunity for the newborn because IgG can cross the placenta Fixing complement Coating antigen for enhanced phagocytosis (Opsonization) Neutralizing toxins and virus Participating in agglutination and precipitating reactions

Types of imMunoglobulin IgM Structure:  IgM normally exists as a pentamer (19S immunoglobulin) but it can also exist as a monomer. In the pentameric form all heavy chains are identical and all light chains are identical. Thus, the valence is theoretically 10. IgM has an extra domain on the mu chain (CH4) and it has another protein covalently bound via a S-S bond called the Joining chain or J chain. This chain functions in polymerization of the molecule into a pentamer. macroglobulin

functions 1. Complement fixation 2. Agglutination 3. Opsonization 4. Toxin Neutralization

Types of imMunoglobulin Properties: a) IgM is the third most common serum Ig. b) IgM is the first Ig to be made by the fetus and the first Ig to be made by a virgin B cells when it is stimulated by antigen. c) As a consequence of its pentameric structure, IgM is a good complement fixing Ig. Thus, IgM antibodies are very efficient in leading to the lysis of microorganisms. d) As a consequence of its structure, IgM is also a good agglutinating Ig . Thus, IgM antibodies are very good in clumping microorganisms for eventual elimination from the body. e) IgM binds to some cells via Fc receptors.

Types of imMunoglobulin f) B cell surface Ig Surface IgM exists as a monomer and lacks J chain but it has an extra 20 amino acids at the C-terminus to anchor it into the membrane. Cell surface IgM functions as a receptor for antigen on B cells. Surface IgM is noncovalently associated with two additional proteins in the membrane of the B cell called Ig-alpha and Ig-beta. These additional proteins act as signal transducing molecules since the cytoplasmic tail of the Ig molecule itself is too short to transduce a signal. Contact between surface immunoglobulin and an antigen is required before a signal can be transduced by the Ig-alpha and Ig-beta chains. In the case of T-independent antigens, contact between the antigen and surface immunoglobulin is sufficient to activate B cells to differentiate into antibody secreting plasma cells. However, for T-dependent antigens, a second signal provided by helper T cells is required before B cells are activated.

Types of imMunoglobulin IgA Structure: Dimer When IgA is found in secretions is also has another protein associated with it called the secretory piece or T piece; sIgA is sometimes referred to as 11S immunoglobulin. Unlike the remainder of the IgA which is made in the plasma cell, the secretory piece is made in epithelial cells and is added to the IgA as it passes into the secretions). The secretory piece helps IgA to be transported across mucosa and also protects it from degradation in the secretions.

Subclasses a) IgA1 - Alpha 1 heavy chains predominant IgA subclass found in serum. Most lymphoid tissues have a predominance of IgA-producing cells. b) IgA2 - Alpha 2 heavy chains the heavy and light chains are not linked with disulfide, but with noncovalent bonds. In secretory lymphoid tissues (e.g., gut-associated lymphoid tissue, or GALT), the share of IgA2 production is larger than in the non-secretory lymphoid organs

Types of imMunoglobulin Secretory Component (SC) Attached to 𝐹 𝐶 region around the hinge portion of alpha chains Consists of 5 Ig-like domains Derived from epithelial cells found in close proximity to the plasma cells Molecular weight of about 70,000

Types of imMunoglobulin Properties: a) IgA is the 2nd most common serum Ig. b) IgA is the major class of Ig in secretions - tears, saliva, colostrum, mucus. Since it is found in secretions secretory IgA is important in local (mucosal) immunity. c) Normally IgA does not fix complement, unless aggregated. d) IgA can binding to some cells - PMN's and some lymphocytes.

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