Chai K X, Ma J X, Murray S R, Chao J, Chao L
Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston 29425.
J Biol Chem. 1991 Aug 25;266(24):16029-36.
The gene encoding rat kallikrein-binding protein (RKBP), a serine protease inhibitor, has been isolated and analyzed with the aid of the polymerase chain reaction. The gene is approximately 10 kilobases in length with four introns of approximately 2.2, 1.8, 0.9, and 0.84 kilobases. This gene is composed of five exons and encodes a polypeptide of 416 amino acid residues. The reactive center region of RKBP is encoded by the fifth exon with the putative P1-P1' residues being Lys-Ser. The organization of the RKBP gene is homologous to those of human alpha 1-antitrypsin and alpha 1-antichymotrypsin in size and arrangement of exons and introns, suggesting that they belong to the same subgroup of serpins. In the 5'-flanking region of the RKBP gene, a variant TATA box sequence, ATAAATA, is found 20 base pairs upstream from the transcription initiation site. The 5'-flanking region of the RKBP gene was able to direct transcription of the reporter gene chloramphenicol acetyltransferase when transfected into a rat hepatoma cell line. An internal promoter-like region was found in the first intron of the RKBP gene, downstream from the transcription initiation site and upstream from the translation initiation codon, however, it was unable to direct expression of the chloramphenicol acetyltransferase reporter gene in our experiments. The expression of RKBP in rat liver was induced by sex hormone treatment as indicated by dot-blot analysis. A genomic Southern blot using an RKBP cDNA probe revealed multiple bands suggesting that the RKBP gene belongs to a family of highly conserved genes.
编码大鼠激肽释放酶结合蛋白（RKBP，一种丝氨酸蛋白酶抑制剂）的基因已借助聚合酶链反应被分离和分析。该基因长度约为10千碱基，有四个内含子，长度分别约为2.2、1.8、0.9和0.84千碱基。此基因由五个外显子组成，编码一个含416个氨基酸残基的多肽。RKBP的反应中心区域由第五个外显子编码，推测的P1 - P1'残基为赖氨酸 - 丝氨酸。RKBP基因的结构在外显子和内含子的大小及排列上与人类α1 - 抗胰蛋白酶和α1 - 抗糜蛋白酶的基因结构同源，表明它们属于丝氨酸蛋白酶抑制剂的同一亚组。在RKBP基因的5'侧翼区域，在转录起始位点上游20个碱基对处发现了一个变异的TATA盒序列ATAAATA。当将RKBP基因的5'侧翼区域转染到大鼠肝癌细胞系中时，它能够指导报告基因氯霉素乙酰转移酶的转录。在RKBP基因的第一个内含子中，在转录起始位点下游和翻译起始密码子上游发现了一个类似内部启动子的区域，然而，在我们的实验中它无法指导氯霉素乙酰转移酶报告基因的表达。斑点印迹分析表明，性激素处理可诱导大鼠肝脏中RKBP的表达。使用RKBP cDNA探针进行的基因组Southern印迹显示出多条带，表明RKBP基因属于一个高度保守的基因家族。
