摘要
用基因重组及定位突变技术成功地构建了t-PA的K1区缺失突变体t-PAdelK1、PAI-1结合位点缺失突变体t-PA del(296—302)及两者的组合突变体t-PA del(K1,296—302),并在COS-7细胞中实现三者的暂时性表达,在CHO细胞中实现了t-PA del(K1,296—302)的稳定性表达。对表达产物的生物学特性分析表明,t-PA del(296—302)及t-PA del(K1,296—302)获得了PAI-1抗性,因时t-PA del(K1,296—302)在大鼠体内的半衰期延长约5倍,而与纤维蛋白的亲和力只略有下降。因此,此组合突变体有可能成为优于野生t-PA的新型溶栓剂候选株。
In this paper, three t-PA mutants, t-PA del K1 (with deletion of Kl domain), t-PA del (296 - 302) (with deletion of,PAI-1 binding site and their combination mutant t-PA del (K1, 296 - 302), were constructed by DNA recombination and site-directed mutagenesis techniques. Then the three t-PA mutants were transiently expressed in COS-7 cells, and the combination mutant t-PA del (K1296-302) was stably expressed in CHO cells. The biological analyses of the expression products demonstrated that t-PA del (296 - 302) and t-PA del (K1, 296 - 302) had obtained the resistance to inhibition by PAI-1. In addition , the half-life of t-PA del (K1, 296 - 302) in rat plasma was increased 5 times while the mutant affinity for fibrin was just a little affected. Therefore, it was reasonable to consider that the mutant t-PA del (K1, 296 - 302) may become a potent candidate of new thrombolytic agent.
出处
《生物工程学报》
CAS
CSCD
北大核心
1995年第1期13-19,共7页
Chinese Journal of Biotechnology
