人纤溶酶原饼环区5基因的原核表达及活性测定

Expression and Its Activity of hPgnK5 Protein in Prokaryotic System

人纤溶酶原饼环区 5 (hPgnK5 )是新的血管生成抑制因子 .PCR法改造hPgnK5基因 ,所得hPgnK5基因包括编码人纤溶酶原C4 62 到P54 4共 83个氨基酸残基 ,而且在 5′ ,3′分别引入了EcoRⅠ和BamHⅠ位点 .以pThiohisA构建hPgnK5基因原核表达载体 ,在大肠杆菌TOP10中表达该蛋白 .通过柱层析法获得hPgnK5纯化蛋白 .免疫印迹反应表明该蛋白具有hPgnK5免疫活性 .体外实验表明 ,该蛋白具有抑制血管内皮细胞增殖活性 .

The human plasminogen kringle 5 is a novel angiogenesis inhibitor.The hPgnK5 gene encoding 83 amino acid residues from C 462 to P 544 was obtained. Eco RⅠ and Bam HⅠ cleavage sites were added to its 5′ and 3′ termini by PCR.The constructed pThiohisA expression vector expressed the hPgnK5 protein in E.coli TOP10.The hPgnK5 protein was purified by column chomatography.Western blot analysis showed that hPgnK5 protein was recognized by mAb against native hPgnK5 and inhibited the proliferation of endothelial cell in vitro. The result shows that the hPgnK5 may be promising in application.