人源孤儿G蛋白偶联受体hGPCRc的分子克隆及其初步鉴定

孤儿G蛋白偶联受体 (orphanGprotein coupledreceptors ,oGPCRs)是最重要的潜在药物靶点 ,对于创新药物研究意义重大 .根据已有文献及相关基因数据库提供的信息 ,利用RT PCR从人结肠组织获得oGPCR某一成员的氨基酸编码序列 ,大小为 10 14bp ,而且与GenBank已登录序列(AB0 835 98)完全一致 ,称之为hGPCRc ;又用相同的引物以健康志愿者血液基因组DNA作为模板进行PCR扩增 ,亦得到同样大小的DNA序列 ,测序显示二者个别碱基不一致 ,但所对应氨基酸序列并无差异 .另外 ,RT PCR对人源部分组织及细胞系的检测结果显示 :hGPCRc在人脑组织表达最高 ,结肠次之 ,其它组织或细胞系如胃、血液、肝、肺、上皮未检测到该基因的表达 .利用相关软件对hGPCRc分别结果显示 :hGPCRc定位于人染色体 13q32 3,与小鼠、大鼠的对应物序列同源性高达85 % ,但与人源其他已知基因的同源性较低 ,对应的氨基酸序列组成了 7个跨膜区段的结构域 .因此 ,hGPCRc符合GPCR的结构特点 ,应为人类oGPCRs的新成员 .

Molecular Cloning and Primary Identification of hGPCRc: a Human Orphan G-protein-coupled Receptor

Orphan G protein-coupled receptors (oGPCRs)are under intense scrutiny as potential targets of drug research.With the aid of the information from gene bank,the whole ORF of a member of oGPCR was amplified by RT-PCR from human colon tissue, and the ORF sequence of the oGPCR,designated hGPCRc,is identical to that in GenBank (AB083598).Also,the ORF of hGPCRc could be obtained with PCR from the template of genomic DNA of a health volunteer's blood.The RT-PCR analysis of different human tissues or cell lines indicated that hGPCRc is expressed most abundantly in brain,less in colon and little in other tissues.Software analysis at the NCBI website showed that hGPCRc was localized at chromosome 13q32.2,and its deduced amino acids formed seven-transmembrane domains and had 85% homology with the counterpart of rat or mouse,but a lower homology with other human genes.Therefore, it was indicated that hGPCRc should be a new member of human oGPCRs.