A novel photoaffinity ligand for the phencyclidine site of the N-methyl-D-aspartate receptor labels a Mr 120,000 polypeptide

• Published in: The Journal of biological chemistry 1990-04, Vol.265 (12), p.6776-6781
• Main Authors: SONDERS, M. S, BARMETTLER, P, LEE, J. A, KITAHARA, Y, KEANA, J. F. W, WEBER, E
• Format: Article
• Language: English
• Subjects: 550201 - Biochemistry- Tracer Techniques; AFFINITY; Affinity Labels - metabolism; AMINES; AMINO ACIDS; ANIMALS; ASPARTIC ACID; Azides - chemical synthesis; Azides - metabolism; AZINES; BASIC BIOLOGICAL SCIENCES; Binding Sites; Binding, Competitive; BIOCHEMICAL REACTION KINETICS; Biological and medical sciences; BODY; BRAIN; Brain - metabolism; CARBOXYLIC ACIDS; CELL CONSTITUENTS; Cell Membrane - metabolism; CELL MEMBRANES; Cell receptors; Cell structures and functions; CENTRAL NERVOUS SYSTEM; Dibenzocycloheptenes - chemical synthesis; Dibenzocycloheptenes - metabolism; Dizocilpine Maleate - analogs & derivatives; ELECTROPHORESIS; Fundamental and applied biological sciences. Psychology; GUINEA PIGS; HETEROCYCLIC COMPOUNDS; HYDROGEN COMPOUNDS; ISOTOPE APPLICATIONS; KINETICS; LIGANDS; Macromolecular Substances; MAMMALS; MEMBRANE PROTEINS; MEMBRANES; Molecular and cellular biology; MOLECULAR WEIGHT; NERVOUS SYSTEM; ORGANIC ACIDS; ORGANIC COMPOUNDS; ORGANIC NITROGEN COMPOUNDS; ORGANS; PEPTIDES; Phencyclidine - metabolism; PIPERIDINES; POLYPEPTIDES; PROTEINS; PYRIDINES; REACTION KINETICS; RECEPTORS; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter - isolation & purification; Receptors, Neurotransmitter - metabolism; Receptors, Neurotransmitter - radiation effects; RODENTS; TRACER TECHNIQUES; TRITIUM COMPOUNDS; Ultraviolet Rays; VERTEBRATES
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(19)39216-6

A radiolabeled photoaffinity ligand has been developed for the N-methyl-D-aspartate (NMDA)-preferring excitatory amino acid receptor complex. [3H]3-Azido-(5S, 10R)(+)-5-methyl-10,11-dihydro-5H- dibenzo[a,d]cyclohepten-5,10-imine [3H]3-azido-MK-801 demonstrated nearly identical affinity, density of binding sites, selectivity, pH sensitivity, and pharmacological profile in reversible binding assays with guinea pig brain homogenates to those displayed by its parent compound, MK-801. When employed in a photo-labeling protocol designed to optimize specific incorporation, [3H]3-azido-MK-801 labeled a single protein band which migrated in sodium dodecyl sulfate-polyacrylamide gels with Mr = 120,000. Incorporation of tritium into this band was completely inhibited when homogenates and [3H]3-azido-MK-801 were coincubated with 10 microM phencyclidine. These data suggest that the phencyclidine site of the NMDA receptor complex is at least in part comprised of a Mr = 120,000 polypeptide.