Search

The NMDA subtype of excitatory glutamate receptor is a multisubunit, fast-acting, ligand-gated cation channel with a high permeability for Ca2+. NMDA receptors have been shown to be of importance in physiological processes, such as long-term potentiation, and also in pathophysiological conditions, including chronic neuronal degeneration and acute excitotoxity (1). Molecular cloning has identified five genes encoding two types of NMDA receptor subunits, the NR1 and NR2A-D subunits. Additionally, the NR1 subunit gene undergoes alternative splicing to yield eight variant forms, thus providing further heterogeneity. The quaternary structures of NMDA receptors are not known, but most are believed to be heteromeric complexes comprising multiple copies of both NR1 and NR2 subunits in as-yet unknown stoichiometries (summarized in 1). Since the absolute polypeptide compositions of native NMDA receptors are unknown, a convenient model system in which to study the properties of defined cloned NMDA receptor subtypes and thus to compare them with their in vivo counterparts is their expression in mammalian cells. This permits the pharmacological, functional and biochemical characterization of either a single type of NMDA receptor subunit or coexpression of combinations of NR1 and NR2 subunit genes (e.g., 2-4). [ABSTRACT FROM AUTHOR]

Antibodies have been widely used in biochemistry as selective probes to study the structure and function of important biological macromolecules. The usefulness of antibody molecules as research tools lies in their unique structure. Antibody molecules consist of two distinct domains; the Fab or "business" end of the molecule, which binds to the respective antigen with its exquisite specificity and the Fc region, which can be readily labeled with a wide range of tags. Prior to the advent of monoclonal antibody technology and the availability of protein primary structures via cDNA cloning, the generation of an antibody against a particular protein required the laborious prerequisite task of antigen purification to homogeneity followed by polyclonal antibody production. Neurotransmrtter receptors are, however, expressed at very low concentrations in mammalian brain, i.e., fmol-pmol receptors/ mg protein, and it was difficult to purify sufficient material for the generation of useful antibodies. However, the recent availability of neurotransmitter receptor cDNAs has revolutionized the generation of antireceptor antibodies worldwide in both academic and commercial laboratories. Receptor cDNAs can be readily manipulated to generate large amounts of protein or parts of the protein of interest, which can then be purified for antigen production. In addition, using the primary structure deduced from the respective cDNA sequences, synthetic oligopeptides can be used as immunogens to generate amino acid sequence-directed antireceptor antibodies. [ABSTRACT FROM AUTHOR]