Structural Biochemistry/Proteins/Antibody and Antigen Interaction

An antibody (also known as immunoglobulin or Ig) is one of an animal's defense mechanisms. One type of antibody, Immunoglobulin G or IgG, has a Y-like structure consisting of two heavy chains and two light chains. The structures of antibodies are partially maintained by disulfide bonds, which link the heavy chains to heavy chains or heavy chains to light chains. The heavy chains always have the same sequence of amino acids while the light chains have varied sequences of amino acids. This variation is important to ensure that the antibody will be able to respond to various antigens, foreign substances that enter animal bodies (e.g.: foreign protein, polysaccharides, nucleic acids, etc.). Antibodies are produced by the B cell, which is a type of white blood cell. B cells will allow the immune system to remember and react faster for future exposures. There are various types of antibodies, which are categorized based on the isotopes that the chains possess. Approximately five different types are discovered in mammals which each carry a special function and response based on the different types of bacteria that they encounter throughout the body. Each of the classes has identical light chains which is matched with a different heavy chain. Each of these classes of immunoglobulin has a specific function. Immunoglobulin M (IgM) is the first class of antibody to appear after exposure to an antigen. IgM has ten combining sites which enables it to bind extra well with antigens containing multiple identical epitopes. An example of avidity, which is the strength of an interaction that contains multiple independent binding interactions between matching antibody-antigen complex. Immunoglobulin A (IgA) is the antibody that is involved in external secretions - tears and saliva - meaning it is the body's first line of defense against bacteria and viruses. The role of Immunoglobulin D (IgD) is still unknown. Immunoglobulin E (IgE) helps to protect against parasites. However, it also participates in allergic reactions. The release of granules containing pharmacologically active molecules is triggered when the IgE - antigen complex forms cross-links with receptors on the surface of mast cells. These antibodies are generated randomly using different gene segments which give a specific bind site for the antibody. They are also randomized by the mutations in the genes, making them even more complex and diverse. This relates to the humoral immune system (see below) that helps humans protect against the wide range of diversity in bacterias and viruses. There are two forms of antibodies: one that is soluble in the blood and fluids of the body and the other one that is attached to the B cell in general. The structure of the antibody includes a heavy chain and a light chain. The heavy chain has a constant region and a variable region and it is about 110 amino acids long. Antibodies first bind to the pathogens to prevent them from damaging cells. Second, they will try to remove pathogens using macrophages. Finally, they will stimulate other immune responses to further destroy the pathogens. The cells and proteins connected with this process are discussed below.

Immune Cell Response- Cells and Proteins Involved

The immune response system is composed of two corresponding systems, the humoral and cellular immune systems. The humoral immune system occurs primarily in body fluids, targeting bacterial infections and extracellular viruses. The humoral system can also react to individual foreign proteins. The cellular immune system attacks host cells infected by viruses as well as some parasite and foreign tissues. Both of these systems are prompted by a category of white blood cells called leukocytes, of which include macrophages (cells that ingest by phagocytosis) and lymphocytes (cells that release antibodies). In the cellular immune system, a class of T cells, called cytotoxic T cells, are the main cells involved with reception of foreign cells or parasites. These cells have T cell receptors that are on the surface of cells and extend through the plasma membrane. When an antigen is deteected, either a T cell receptor or antibody will bind to a specific molecular structure in the antigen. This structure is referred to as antigenic determinant or an epitope (see Epitopes under Protein Function). Helper T cells are also a part of the cellular immune system, generating cytokines, a type of soluble signaling proteins. In clonal selection, helper T cells are involved only indirectly, prompting the selective reproduction of the cytotoxic T cells and B cells that can bind to the certain antigen.

In performing this response, antibodies can either be:
 * Monoclonal
 * One type of antibody responding to one type of antigen by recognizing one antigenic determinant.


 * Polyclonal
 * Several type of antibody responding to one type of antigen by cooperatively recognizing various antigenic determinants on the antigen.

Epitopes

Epitopes are the sites on a antigen that the immune system recognizes. It is also called an antigenic determinant. Both the host and foreign protein can produce epitopes that can bind to the paratopes of the B and T lymphocytes. An epitope can be either linear or conformational epitope. Linear epitotes are recognized by long chain of amino acids, or their primary structure: the sequence of the animo acids. Conformational epitopes are recognized by the antibodies by the 3-D structures of the epitopes.

Antibody Folding

Recent studies have shown how antibodies fold. Foldings occur in the endoplasmic reticulum even before chains complete translation. Most of these studies are made by dissecting the antibody and letting it denaturalize and refold. IgGs are mostly used for these experiments. IgGs mostly form through adding a light chain to a heavy chain dimer between the CL and CH1 region. Dissection of IgGs light chain shows that there are three pathways of folding caused and limited to proline cis-trans isomerization. In general, most antibodies are formed through these three pathways after the C2-C3 sulfide bridge formed.

In equilibrium, most proteins have the accessible conformations, such as the native state, unfolded state, and non specific aggregates. However, At low pH (pH<3), Antibodies have tendency to adopt specific additional conformation, called “alternating folded state”( though most other proteins are unfolded in this environment). Through spectroscopic, the structure is different from the nature state. However, it exhibits characteristic of the folded state ( e.g. stability against unfolding). The concern about this process is significant in biotechnology because antibody manufacturing process often include low pH steps, which can easily perform the “alternatively folded state”. For some antibodies, another accessible state is the fibrillar amyloid structure, that can lead to some protein-folding diseases. Fibrillar amyloid structure is a cross-beta structure in which fibrils are formed by beta-strand exchange of the individual subunits. Isolated LCs and truncated HCs form fibrils and are deposited in organs such as kidneys. The large quantity deposit can cause some fatal diseases when it interferes with physiological functions. E.g. A large deposit of monoclonal LCs prone to misfolding and the formation of amyloid deposit can cause a fatal disease [light chain amyloidosis (AL)].The fibrilization mechanism is still under study. However, the mechanism in production of variable domains might generate less stable domain, therefore, can pass ER quality control but have a propensity to misfold outside the cell. B cell: a stage in which heavy and light chains are synthesized and expressed at cell surface via a transmembrance. Heavy/light chain: constituent polypeptide chains of antibody molecules. Light chains are made up of two Ig domains. Heavy chains are made up of minimum of 3 Ig domains. Ig domain: 100 amino acid-folding unit conserves twisted barrel-like-beta-sheet structure. It is stabilized by a buried intrachain disulfide bond.

Clonal Expansion

When the antibodies recognizes a specific antigen, the B cells clone itself and turn into 2 types of cells: the plasma cells and the memory cells. The plasma cells are effector cells that produce a large amount of a specific soluble antibody that attacks the target antigen. Memory Cells are long lived cells that can be quickly activated to produce antibodies when the specific antigen is observed.

During the primary Immune response, that is the first time antibody attacks an antigen, it takes a long time for the antibody to make a large amount of plasma cells and memory cells when compared to the secondary response. During the secondary immune response, there is only a short time lag and the rate of Antibody produced is a lot greater. This is due to the memory cells activating the production of antibodies.

Reference
Nelson, David L., and Michael M. Cox. Principles of Biochemistry. 5th ed. New York: W.H Freeman and Company, 2008. Buchner, Johannes; Feige, Matthias J.; and Hendershot, Linda M. "How antibodies fold". Trends in Biochemical Sciences. Vol. 5, (4). doi:10.1016/j.tibs.2009.11.005.