科研之友
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摘要
A gamma-aminobutyric acid, (GABA(A)) receptor (GABA(A)R) gamma, subunit (short form) was cloned from an adult human cerebral cortex cDNA library in bacteriophage lambdagt11. The 26 1 -bp intracellular loop (IL) located between M3 and M4 was amplified using the polymerase chain reaction and inserted into the expression vectors lambdagt11 and pGEX-3X. Both beta-galactosidase (LacZ) and glutathione-S-transferase (GST) fusion proteins containing the gamma2IL were purified, and a rabbit antibody to the LacZ-gamma2IL was made. The antibody reacted with the gamma2IL of both LacZ and GST fusion proteins and immunoprecipitated the GABA(A)R/benzodiazepine receptor (GABA(A)R/BZDR) from bovine and rat brain. The antibody reacted in affinity-purified GABA(A)R/BZDR immunoblots with a wide peptide band of 44,000-49,000 M(r). Immunoprecipitation studies with the anti-gamma2IL antibody suggest that in the cerebral cortex, 87% of the GABA(A)Rs with high affinity for benzodiazepines and 70% of the GABA(A)Rs with high affinity for muscimol contain at least a gamma subunit, probably a gamma2. These results indicate that there are [H-3]muscimol binding GABA(A)Rs that do not bind [H-3]flunitrazepam with high affinity. Immunoprecipitations with this and other anti-GABA(A)R/BZDR antibodies indicate that the most abundant combination of GABA(A)R subunits in the cerebral cortex involves alpha1, gamma2 (or other gamma), and beta2 and/or beta3 Subunits. These subunits coexist in >60% of the GABA(A)R/BZDRs in the cerebral cortex. The results also show that a considerable proportion (20-25%) of the cerebellar GABA(A)R/BZDRs is clonazepam insensitive. At least 74% of these cerebellar receptors, which likely contain alpha6, also contain gamma2 (or other gamma) subunit(s). The alpha1 and beta2 or beta3 subunits are also frequently associated with gamma2 (or other gamma) and alpha6 in these cerebellar receptors.
- 出版日期1993-3
