JOINT STUDY ON STRUCTURE AND FUNCTION OF GABA_B RECEPTOR

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We have previously reported that a GABA_B receptor, approximately 80 kDa in size, purified from the bovine cerebral cortex, is characterized to be one of G protein-coupled receptors. Recently, we obtained a partial amino acid sequence of the purified receptor protein, and its deduced DNA sequences were utilized for synthesizing PCR primers in various combinations of antisense oligonucleotides. In addition, another set of PCR primers was designed to include one of sequences which have been found commonly in transmembrane domains of various G protein-coupled receptors. Using various combinations of these two types of primers, PCR amplification of mRNA from the rat cerebellum gave us a PCR product. Its partial sequence indicated that the cDNA of the product appeared to code a protein possessing hydrophobic, probably transmembrane, regions. We therefore tried to extend this cDNA product toward both 5'-and 3'-ends by using the method of rapid amplification of cDNA ends (RACE). Our analysis of the RACE-products indicated that the sequences of both nucleotide and their deduced amino acid possessed possible transmembrane domains which were highly homologous to those of certain G protein-coupled receptors. Our further analysis by RT-PCR revealed that the RACE-products were expressed not only in the cerebellum but also in the forebrain, midbrain, pons, medulla and spinal cord of the rat. The present study suggests that the PCR product reported here may represents the presence of a novel G protein-coupled protein.