Immunoaffinity purification of GABAA receptor alpha-subunit iso-oligomers. Demonstration of receptor populations containing alpha 1 alpha 2, alpha 1 alpha 3, and alpha 2 alpha 3 subunit pairs.

Duggan, M.J., Pollard, S. and Stephenson, F.A. (1991) Immunoaffinity purification of GABAA receptor alpha-subunit iso-oligomers. Demonstration of receptor populations containing alpha 1 alpha 2, alpha 1 alpha 3, and alpha 2 alpha 3 subunit pairs. Journal of Biological Chemistry, 266 (36). pp. 24778-24784.

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Abstract

Novel methods for the isolation of gamma-aminobutyric acidA (GABAA) receptor alpha subunit iso-oligomers have been developed. Thus, populations of GABAA receptors containing the GABAA receptor alpha 1 subunit, the alpha 2 subunit, and the alpha 3 subunit have been purified from sodium deoxycholate extracts of bovine cerebral cortex with the retention of specific [3H]flunitrazepam-binding activity by anti-alpha 1 324-341, anti-Cys alpha 2 414-424, or anti-Cys alpha 3 454-467 antibody affinity chromatography, respectively. The relative abundance of the different specificity alpha subunits in these preparations was compared with benzodiazepine affinity chromatography-purified GABAA receptors by immunoblotting. In each case, it was found that although the immunoreactivity with the specific alpha subunit antibody that was used for purification was enriched in immunoaffinity-purified receptors, reactivity with the other alpha subunit specificity antibodies, together with anti-gamma 2 1-14 Cys immunoreactivity was found. Immunoprecipitation of GABAA receptors purified by anti-alpha 1 324-341 antibody affinity chromatography by all three anti-alpha subunit antibodies employed, together with the use of anti-alpha 1 324-341 and anti-Cys alpha 2 414-424 antibody affinity columns in series, further substantiated the partial co-purification of the different polypeptides. These results demonstrate the copurification of the gamma 2 subunit with each population of alpha 1, alpha 2, alpha 3 subunit-enriched GABAA receptors. They also show the existence of minor populations of GABAA receptors that contain alpha 1 alpha 2, alpha 1 alpha 3, and alpha 2 alpha 3 subunit pairs within single oligomers.

Item Type:Article
Departments, units and centres:Department of Pharmaceutical and Biological Chemistry > Department of Pharmaceutical and Biological Chemistry
ID Code:2581
Journal or Publication Title:Journal of Biological Chemistry
Deposited By:Library Staff
Deposited On:25 Nov 2011 16:22
Last Modified:25 Nov 2011 16:22

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