RGD肽与尿激酶B链融合蛋白原核重组表达质粒的构建,表达和？…

将化学合成的ＲＧＤ肽（Ａｒｇ－Ｇｌｙ－Ａｓｐ）编码寡核苷酸与尿激酶Ｂ链ｃＤＮＡ相连成为融合基因后，克隆至原核表达质粒ｐＢＶ２２０中，在ＰＲＰＬ自动子的作用下，经４２℃热诱导，在大肠杆菌ＤＨ５α中获得了融合基因的表达，其表达量占菌体总蛋白的９．２％，表达产物以无活性的包含体形式存在。经变复性处理得到纯化的融合基因的表达产物，经Ｗｅｓｔｅｒｎ－ｂｌｏｔｔｉｎｇ分析表明产物具有与天然尿激酶相似的抗原性，

Construction of Fusion Recombinant Plasmid Coding for RGD Peptide and Urokinase B Chain, Its Expression in Escherichia coil and Preliminary Characterization of Its Biological Activity

A synthetic RGD (Arg-Gly-Asp) peptide-coding sequence was fused with urokinase B-chain cDNA and then cloned into the prokaryotic expression vector pBV220. The fused gene was expressed in E. coli DH5 alpha under the control of PRPL promoter by 42 degrees C induction. The expression level of the fusion protein was over 9.2% of the total bacterial proteins as a or of inactive inclusion body. The purified fusion protein was obtained with similar antigenicity as urokinase shown by Western blotting. Its in vitro fibrinolysis and anti-platelet aggregation activity was also evaluated by bioassay.