毕赤酵母重组Annexin32的分子特征及其对血凝与血栓形成的影响

Characteristics of the Recombinant Annexin 32 in Pichia pastoris and its Effects on Coagulation Time and Thrombosis

采用SDS PAGE、等电聚焦、脱磷酸反应、蛋白质N端氨基酸测序、KPTT及大鼠下腔静脉血栓形成观察 ,发现毕赤酵母重组Annexin32的分子特征较原核产物有了明显变化。有主、次 2条带 ,主带的分子量比原核产物大 ,等电点较理论值低 ,已被磷酸化 ,2条带的N端均携带了来自载体的 4个氨基酸 ,可能对其与活化血小板表面磷脂的结合有所下调 ,导致血凝与血栓形成受到一定的影响 ,但仍具有明显的抗凝及抑制血栓形成作用。

The recombinant annexin32 in Pichia pastoris was assayed by SDS-PAGE,IEF,dephosphorization,sequencing of amino-N-terminal,KPTT and venous thrombosis in inferior vena cava of rats.Characteristics of the recombinant annexin32 in Pichia pastoris varied from that of in E.coli. The r-annexin32 showed two bands.MW of the majar band was larger than that of r-annexin32 in E.coli.Its pI was lower than the theorial pI value of annexin32 because of phosphorization.The amino-N-terminus of two bands carried 4 aminoes from the yeast expression plasmid.These changes might decrease affinity with phospholipid-binding to activated pellet and affect coagulation time and thrombosis.However,the r-annexin32 in Pichia pastoris still had strong anticoagulation and antithrombosis.;