摘要
Three tissue-type plasminogen activator(t-PA)mutants were constructed by recombinant andsite-directed mutagenesis techniques.They are del(296—302)with deletion of PAI-1 binding site,N117Q/N184Qwith deglycosylation of K1 and K2 domains,and their combination mutant designated as GGI.Then these threemutants were suocessfully transiently expressed in COS-7 ceils,and GGI was further stably expressed in CHOcells.The biological characterization of the expression products indicated that del(296—302)and GGIpossessed the resistance to inhibition by PAI-1.In addition,the specific activity of GGI was increased byabout 46,the plasma half-life was prolonged by about one fold,while its affinity for fibrin was not affected.
Three tissue-type plasminogen activator(t-PA)mutants were constructed by recombinant and site-directed mutagenesis techniques.They are del(296—302)with deletion of PAI-1 binding site,N117Q/N184Q with deglycosylation of K1 and K2 domains,and their combination mutant designated as GGI.Then these three mutants were suocessfully transiently expressed in COS-7 ceils,and GGI was further stably expressed in CHO cells.The biological characterization of the expression products indicated that del(296—302)and GGI possessed the resistance to inhibition by PAI-1.In addition,the specific activity of GGI was increased by about 46,the plasma half-life was prolonged by about one fold,while its affinity for fibrin was not affected.
