Biochemical evidence for the existence of gamma-aminobutyrateA receptor iso-oligomers

• Published in: The Journal of biological chemistry Vol. 265; no. 7; pp. 3831 - 3835
• Main Authors: Duggan, M J, Stephenson, F A
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 05.03.1990
• Subjects: Amino Acid Sequence; Animals; Antibodies - isolation & purification; Binding Sites; Brain - metabolism; Cattle; Flunitrazepam - metabolism; Molecular Sequence Data; Organ Specificity; Peptide Fragments - isolation & purification; Receptors, GABA-A - genetics; Receptors, GABA-A - immunology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(19)39669-3

Polyclonal antibodies were raised against synthetic peptides whose sequences were from unique regions of the bovine gamma-aminobutyrateA receptor alpha 1, alpha 2, and alpha 3 subunits. The anti-alpha 1 324-341, anti-Cys alpha 2 414-424, and anti-Cys alpha 3 454-467 antibodies all specifically immunoprecipitated [3H]flunitrazepam and [3H]muscimol binding activities in parallel from Na+ deoxycholate extracts of bovine cerebral cortex. The maximum number of benzodiazepine binding sites immunoprecipitated by each antibody in three brain regions, cerebral cortex, cerebellum, and hippocampus, was investigated. Differences were found for both the maximum number of sites immunoprecipitated by each antibody in one brain region and for the percentage of benzodiazepine binding sites immunoprecipitated by one specificity antibody between the different brain regions. Furthermore, it was found that co-immunoprecipitation with either anti-alpha 1 324-341, anti-Cys alpha 2 414-424, and anti-Cys alpha 3 454-467 or anti-alpha 1 324-341 and anti-Cys alpha 3 454-467 antibodies resulted in an increase in the percentage of benzodiazepine binding sites immunoprecipitated, the sum of which was equal to the percentages pelleted by the individual antibodies. These results demonstrate for the first time the existence in mammalian brain of gamma-aminobutyrateA receptor alpha subunit iso-oligomers.