Solubilization, characterization, and partial purification of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-, quisqualate-, kainate-sensitive L-glutamate binding sites from porcine brain synaptic junctions.

Article date: 1991/12/1

PubMed ID: 1682417

Journal name: Journal of neurochemistry (ISSN: 0022-3042)


L-[3H]Glutamate binding sites were solubilized from porcine brain synaptic junctions by Triton X-114 in the presence of KCl. The solubilized binding sites bound L-[3H]glutamate reversibly with KD and Bmax values of 1.48 +/- 0.18 microM and 178.2 +/- 15.9 pmol/mg of protein, respectively. These binding sites appeared to be integral membrane glycoproteins, with sugar moieties recognized by wheat germ agglutinin. A 49.3-fold purification of these binding sites was achieved by Triton X-114 solubilization, anion-exchange chromatography, and affinity chromatography using wheat germ agglutinin-Sepharose. The apparent molecular mass of the partially purified binding sites was 620 +/- 50 kDa. L-[3H]Glutamate bound to the solubilized preparation could be effectively displaced by agonists of non-N-methyl-D-aspartate (NMDA) L-glutamate receptors but not by NMDA or alpha-amino-4-phosphonobutyrate. The rank order for the competitive ligands in displacing L-[3H]glutamate was: quisqualate greater than alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid greater than L-glutamate greater than kainate.

Author List: Chang Y C, Lin Y H, Lee Y H, Leng C H

Publication Types: Journal Article; Research Support, Non-U.S. Gov't

Substances mentioned in the article: Detergents; Glutamates; Ibotenic Acid; Polyethylene Glycols; Glutamic Acid; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Quisqualic Acid; Nonidet P-40; Kainic Acid;

Mesh terms: Animals; Binding Sites/drug effects; Brain/metabolism; Chromatography; Detergents; Glutamates/metabolism; Glutamic Acid; Hot Temperature; Ibotenic Acid/analogs & derivatives; Kainic Acid/pharmacology; Polyethylene Glycols; Quisqualic Acid/pharmacology; Solubility; Synapses/metabolism; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid;

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