人纤溶酶原Kringle1-5结构域的表达及活性鉴定  

Expression and Characterization of Kringle 1-5 Domains of Human Plasminogen

作　　者：周庆玮[1] 谢静莉[2] 辛利[1] 叶勤[2] 李载平[1] 甘人宝[1] 

机构地区：[1]中国科学院上海生命科学研究院生物化学与细胞生物学研究所,上海200031 [2]华东理工大学生物反应器国家重点实验室,上海200237

出　　处：《生物化学与生物物理学报》2003年第8期761-767,共7页

摘　　要：利用RT PCR的方法从人肝癌细胞株HepG2细胞内获得了编码人纤溶酶原 (hPlasminogen)的Kringle 1到 5(简称K1- 5 )的cDNA ,将其克隆到表达载体pHIL S1中。将重组载体pHIL K1- 5转化毕赤酵母GS115 ,得到的重组菌株用甲醇进行诱导表达 ,并利用赖氨酸亲和柱纯化重组蛋白质。重组蛋白质K1- 5能特异性地按剂量依赖的方式抑制碱性成纤维细胞生长因子 (bFGF)刺激的牛主动脉内皮细胞 (BAEC)的增殖 ,浓度为 14mg L时达到最大抑制效果的 5 0 % ;K1- 5能抑制bFGF引起的BAEC的迁移 ,5 0mg L的K1- 5对BAEC迁移的抑制率为 4 7% ;K1- 5还能影响BAEC细胞的周期 ,14mg L的K1- 5使细胞在G0 ～G1 期积聚。The cDNA encoding Kringle 1-5 domains of human plasminogen (designated as K1-5), obtained from HepG2 by RT PCR, was cloned into expression vector pHIL S1. The recombinant plasmid pHIL K 1 -5 was transformed into Pichia pastoris GS115 and the recombinant yeast was induced by methanol to express the recombinant protein. The expressed protein was purified by lysine affinity chromatography. The recombinant K1-5 inhibited the growth of bovine aortic endothelial cells (BAEC) stimulated by the basic fibroblast growth factor (bFGF), in a dosage dependent manner, with a half maximal concentration of 14 mg/L. And rhK1-5 inhibited 47% of the BAEC migration stimulated by bFGF at the concentration of 50 mg/L. rhK1-5 also affected the cell cycle of BAEC and caused G 0 G 1 arrest at the concentration of 14 mg/L.

关 键 词：人纤溶酶原 结构域 表达 纯化 生物活性 毕赤酵母 

分 类 号：Q78[生物学—分子生物学]