Duggan M J, Stephenson F A
Department of Pharmaceutical Chemistry, School of Pharmacy, London, United Kingdom.
J Biol Chem. 1990 Mar 5;265(7):3831-5.
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.
针对合成肽制备了多克隆抗体，这些合成肽的序列来自牛γ-氨基丁酸A受体α1、α2和α3亚基的独特区域。抗α1 324 - 341、抗半胱氨酸α2 414 - 424和抗半胱氨酸α3 454 - 467抗体均能从牛大脑皮层的Na+脱氧胆酸盐提取物中特异性地共免疫沉淀[3H]氟硝西泮和[3H]蝇蕈醇结合活性。研究了三种脑区（大脑皮层、小脑和海马体）中每种抗体免疫沉淀的苯二氮䓬结合位点的最大数量。发现一个脑区中每种抗体免疫沉淀的位点最大数量以及不同脑区中一种特异性抗体免疫沉淀的苯二氮䓬结合位点百分比均存在差异。此外，还发现用抗α1 324 - 341、抗半胱氨酸α2 414 - 424和抗半胱氨酸α3 454 - 467抗体或抗α1 324 - 341和抗半胱氨酸α3 454 - 467抗体进行共免疫沉淀，会导致免疫沉淀的苯二氮䓬结合位点百分比增加，其总和等于各单独抗体沉淀的百分比。这些结果首次证明了哺乳动物大脑中存在γ-氨基丁酸A受体α亚基异寡聚体。
