N-Methyl-D-aspartate/Phencyclidine Receptor Complex of Rat Forebrain: Purification and Biochemical Characterization

Annat F. Ikin; Yoel Kloog; Mordechai Sokolovsky

doi:10.1021/bi00461a012

N-Methyl-D-aspartate/Phencyclidine Receptor Complex of Rat Forebrain: Purification and Biochemical Characterization

Annat F. Ikin, Yoel Kloog, Mordechai Sokolovsky^*

^*Corresponding author for this work

Department of Neurobiology

Tel Aviv University

Research output: Contribution to journal › Article › peer-review

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Abstract

The N-methyl-D-aspartate (NMDA)/phencyclidine (PCP) receptor from rat forebrain was solubilized with sodium cholate and purified by affinity chromatography on amino-PCP-agarose. A 3700-fold purification was achieved. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and dithiothreitol revealed four major bands of M_r 67000, 57 000, 46000, and 33 000. [³H]Azido-PCP was irreversibly incorporated into each of these bands after UV irradiation. The dissociation constant (K_d) of [1-(2-thienyl)cyclohexyl]piperidine ([³H]TCP) binding to the purified NMDA/PCP receptor was 120 nM. The maximum specific binding (B_max) for [³H]TCP binding was 3.3 nmol/mg of protein. The pharmacological profile of the purified receptor complex was similar to that of the membranal and soluble receptors. The binding of [³H]TCP to the purified receptor was modulated by the NMDA receptor ligands glutamate, glycine, and NMDA.

Original language	English
Pages (from-to)	2290-2295
Number of pages	6
Journal	Biochemistry
Volume	29
Issue number	9
DOIs	https://doi.org/10.1021/bi00461a012
State	Published - 1 Mar 1990

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title = "N-Methyl-D-aspartate/Phencyclidine Receptor Complex of Rat Forebrain: Purification and Biochemical Characterization",

abstract = "The N-methyl-D-aspartate (NMDA)/phencyclidine (PCP) receptor from rat forebrain was solubilized with sodium cholate and purified by affinity chromatography on amino-PCP-agarose. A 3700-fold purification was achieved. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and dithiothreitol revealed four major bands of Mr 67000, 57 000, 46000, and 33 000. [3H]Azido-PCP was irreversibly incorporated into each of these bands after UV irradiation. The dissociation constant (Kd) of [1-(2-thienyl)cyclohexyl]piperidine ([3H]TCP) binding to the purified NMDA/PCP receptor was 120 nM. The maximum specific binding (Bmax) for [3H]TCP binding was 3.3 nmol/mg of protein. The pharmacological profile of the purified receptor complex was similar to that of the membranal and soluble receptors. The binding of [3H]TCP to the purified receptor was modulated by the NMDA receptor ligands glutamate, glycine, and NMDA.",

author = "Ikin, {Annat F.} and Yoel Kloog and Mordechai Sokolovsky",

year = "1990",

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AU - Sokolovsky, Mordechai

PY - 1990/3/1

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N2 - The N-methyl-D-aspartate (NMDA)/phencyclidine (PCP) receptor from rat forebrain was solubilized with sodium cholate and purified by affinity chromatography on amino-PCP-agarose. A 3700-fold purification was achieved. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and dithiothreitol revealed four major bands of Mr 67000, 57 000, 46000, and 33 000. [3H]Azido-PCP was irreversibly incorporated into each of these bands after UV irradiation. The dissociation constant (Kd) of [1-(2-thienyl)cyclohexyl]piperidine ([3H]TCP) binding to the purified NMDA/PCP receptor was 120 nM. The maximum specific binding (Bmax) for [3H]TCP binding was 3.3 nmol/mg of protein. The pharmacological profile of the purified receptor complex was similar to that of the membranal and soluble receptors. The binding of [3H]TCP to the purified receptor was modulated by the NMDA receptor ligands glutamate, glycine, and NMDA.

AB - The N-methyl-D-aspartate (NMDA)/phencyclidine (PCP) receptor from rat forebrain was solubilized with sodium cholate and purified by affinity chromatography on amino-PCP-agarose. A 3700-fold purification was achieved. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and dithiothreitol revealed four major bands of Mr 67000, 57 000, 46000, and 33 000. [3H]Azido-PCP was irreversibly incorporated into each of these bands after UV irradiation. The dissociation constant (Kd) of [1-(2-thienyl)cyclohexyl]piperidine ([3H]TCP) binding to the purified NMDA/PCP receptor was 120 nM. The maximum specific binding (Bmax) for [3H]TCP binding was 3.3 nmol/mg of protein. The pharmacological profile of the purified receptor complex was similar to that of the membranal and soluble receptors. The binding of [3H]TCP to the purified receptor was modulated by the NMDA receptor ligands glutamate, glycine, and NMDA.

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N-Methyl-D-aspartate/Phencyclidine Receptor Complex of Rat Forebrain: Purification and Biochemical Characterization

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