人SBK1 cDNA的克隆及其相互作用蛋白的筛选

首次克隆到人的SBK1（homo sapiens SH3-binding domain kinase 1，SBK1）的cDNA序列，并通过生物信息学的手段，电子克隆到人SBK1的基因组DNA序列.人的SBK1是鼠SBK1的直系同源物，两者基因组DNA结构相似，均含有4个外显子.人的sbk1基因ORF长1 275 bp，编码424个氨基酸，而鼠的ORF长1 254 bp，编码417个氨基酸.两者编码区的核苷酸序列同源性达87.7%，而氨基酸序列同源性达95.7%，在羧基端均有一个PV富集区，推测其能与含有SH3结构域的蛋白质结合.将RT－PCR所获得的长度为1 610 bp的sbk1cDNA序列搜索EST数据库，进行电子延伸，最终获得了约5 kb的人sbk1全长mRNA序列，它与鼠的sbk1全长mRNA大小一致；通过比较基因组学发现UniGene族Hs.97837实际上代表了sbk1基因UniGene族Hs.460471的3′UTR区域，而不是代表了一个新的UniGene族.采用酵母双杂交技术，以SBK1为“诱饵”，获得了与之相互结合的蛋白表皮生长因子受体EGFR和核孤儿受体蛋白NR4A1，它们之间的具体功能关系有待进一步研究.

Cloning of Human SBK1 cDNA and Screening of Its Interaction Proteins

We have cloned a cDNA sequence of the human kinase SBK1(homo sapiens SH3-binding domain kinase 1)for the first time.The genomic DNA sequence of human SBK1 has been acquired through an electronic clone by the bioinformatics tools. Human SBK1 is the orthologue of mouse SBK1.They both have similar genomic DNA structure,and they are composed of four exons.Human sbk1 gene contains an open reading frame(ORF) of 1 275 bp length,encoding 424 amino acids,while the ORF of mouse is 1 254 bp length,encoding 417 amino acids. Both sbk1 genes share 87.7% nucleotide homologue and 95.7% amino acid homologue in the coding region.A common PV amino acids rich region in the carboxyl terminus are assumed to bind a certain SH3-domain containing protein.The 1 610 bp length cDNA sequence of human SBK1 from RT-PCR was performed an electronic extension to gain a full length mRNA sequence about 5 kb by searching the EST database, which is equal to the mRNA size of mouse.Furthermore,with comparative genomics method we found that the UniGene Hs.97837 was actually the 3'UTR region of human sbk1 UniGene Hs.460471, rather than a novel UniGene.A yeast two-hybrid screening was performed with human SBK1 as the bait.As a result,SBK1 can bind EGFR(epidermal growth factor receptor) and NR4A1(nuclear receptor subfamily 4, group A, member 1) in yeast cells,but the functional relationship between SBK1 and EGFR or NR4A1 still needs further investigation.