Purification and pharmacological and immunochemical characterization of synaptic membrane proteins with ligand-binding properties of N-methyl-D-aspartate receptors.

摘要: A method was developed for the solubilization of approximately 50% proteins in synaptic membranes that have ligand-binding characteristics N-methyl-D-aspartate (NMDA) receptors. Affinity chromatographic separation solubilized through L-glutamate-derivatized matrices and subsequent elution by NMDA-containing buffers led to purification four predominant with estimated sizes 67-70, 53-62, 41-43, 28-36 kDa. The co-purification NMDA-sensitive L-glutamate binding, dizocilpine-sensitive thienylcyclohexyl piperidine (TCP)-binding, strychnine-insensitive glycine-binding achieved this affinity procedure. Glutamate, glycine, polyamine spermidine increased both "on" rate equilibrium level [3H]TCP binding isolated proteins. group eluted NMDA from glutamate-derivatized could be further purified size exclusion chromatography glutamate-binding TCP-binding Polyclonal monoclonal antibodies cloned receptor protein NMDAR1 did not react any membrane or fractions. However, immunoreaction raised against a phosphonoaminocarboxylic acid-binding indicated these are two major These studies indicate might components complex has some receptors neuronal may contain varieties NMDA-like composed subunits differ NMDAR2