The base of the Y plays a role in modulating immune cell activity. This region is called the Fc Fragment, crystallizable region, and is composed of two heavy chains that contribute two or three constant domains depending on the class of the antibody. By binding to specific proteins the Fc region ensures that each antibody generates an appropriate immune response for a given antigen.
The Fc region also binds to various cell receptors, such as Fc receptors, and other immune molecules, such as complement proteins. By doing so, it mediates different physiological effects including opsonization, cell lysis, and degranulation of mast cells, basophils and eosinophils.
A single-chain variable fragment scFv is a fusion protein of the variable regions of the heavy VH and light chains VL of immunoglobulins, connected with a short linker peptide of ten to about 25 amino acids. The linker is usually rich in glycine for flexibility, as well as serine or threonine for solubility, and can either connect the N-terminus of the VH with the C-terminus of the VL, or vice versa. This protein retains the specificity of the original immunoglobulin, despite removal of the constant regions and the introduction of the linker.
These molecules were created to facilitate phage display , where it is highly convenient to express the antigen-binding domain as a single peptide. As an alternative, scFv can be created directly from subcloned heavy and light chains derived from a hybridoma. ScFvs have many uses, e. Unlike monoclonal antibodies, which are often produced in mammalian cell cultures, scFvs are more often produced in bacteria cell cultures such as E. The fragment antigen-binding Fab fragment is a region on an antibody that binds to antigens.
It is composed of one constant and one variable domain of each of the heavy and the light chain. The two variable domains bind the epitope on their specific antigens. In an experimental setting, Fc and Fab fragments can be generated in the laboratory. The enzyme papain can be used to cleave an immunoglobulin monomer into two Fab fragments and an Fc fragment. The enzyme pepsin cleaves below hinge region, so a F ab' 2 fragment and a pFc' fragment is formed.
The F ab' 2 fragment can be split into two Fab' fragments by mild reduction. The variable regions of the heavy and light chains can be fused together to form a single-chain variable fragment scFv , which is only half the size of the Fab fragment, yet retains the original specificity of the parent immunoglobulin.
A diabody is two scFvs with connected with linker peptides that are too short for the two variable regions to fold together about five amino acids , forcing the scFvs to dimerize. Diabodies have been shown to have dissociation constants up to fold lower than corresponding scFvs, meaning that they have a much higher affinity to their target. Two scFv's which bind to the same or different antigens may also be connected with longer linkers such as leucine zippers.
Educational Appendix. Antibody Structure Antibodies are glycoproteins. Resistant to digestion and is secreted in milk. Major Ig in serum. Provides the majority antibody based in immunity against invading pathogens. Moderate complement fixer. IgG3 can cross placenta.
The type of heavy chain present defines the class of an antibody. Each heavy chain has two regions, the constant region and the variable region. The constant region is identical in all antibodies of the same isotype, but differs in antibodies of different isotypes. The variable region of the heavy chain differs depending on the B cell that produced it, but is the same for all antibodies produced by a single B cell or B cell clone. The variable region of each heavy chain is approximately amino acids long and is composed of a single Ig domain.
A light chain has two successive domains: one constant domain and one variable domain. The approximate length of a light chain is — amino acids. Each antibody contains two light chains that are always identical. The Y-shape of an antibody can be divided into three sections: two F ab regions and an Fc region. The F ab regions contain the variable domain that binds to cognate antigens. The Fc fragment provides a binding site for endogenous Fc receptors on the surface of lymphocytes, and is also the site of binding for secondary antibodies.
In addition, dye and enzymes can be covalently linked to antibodies on the Fc portion of the antibody for experimental visualization. These three regions can be cleaved into two F ab and one Fc fragments by the proteolytic enzyme pepsin. Antibody fragments have distinct advantages in certain immunochemical techniques. Fragmenting IgG antibodies is sometimes useful because F ab fragments 1 will not precipitate the antigen and 2 will not be bound by immune cells in live studies because of the lack of an Fc region.
Often, because of their smaller size and lack of crosslinking due to loss of the Fc region , F ab fragments are radiolabeled for use in functional studies. Fc fragments are often used as Fc receptor blocking agents in immunohistochemical staining. Return to the antibody guide.
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