CAJ | 학술논문

目的利用甲醇酵母高效表达重组人血小板第4因子(小PF4)。方法用PCR方法扩增出血小板第4因子(PF4)的cDNA序列，插入到穿梭质粒pPIC9的MFa信号肽后，在醇氧化脱氢酶1(AOXI)启动子的下游，得到重组质粒pPIC9-PF4。转化到甲醇酵母宿主菌GS115，经筛选得到PF4高表达菌株进行表达，并测定表达产物氨基酸序列和生物学功能。结果 rhPF4氨基酸序列与天然PF4相同，可中和肝素抗凝作用，并呈剂量相关。结论 rhPF4可在甲醇酵母中获得生产规模表达，产物性质和天然的PF4相同。
목적이용갑순효모고효표체중조인혈소판제4인자(소PF4)。방법용PCR방법확증출혈소판제4인자(PF4)적cDNA서렬，삽입도천사질립pPIC9적MFa신호태후，재순양화탈경매1(AOXI)계동자적하유，득도중조질립pPIC9-PF4。전화도갑순효모숙주균GS115，경사선득도PF4고표체균주진행표체，병측정표체산물안기산서렬화생물학공능。결과 rhPF4안기산서렬여천연PF4상동，가중화간소항응작용，병정제량상관。결론 rhPF4가재갑순효모중획득생산규모표체，산물성질화천연적PF4상동。
Objective To express recombinant human platelet factor 4 (PF4) in methanol yeast. Methods PF4 cDNA amplified by PCR was cloned into the mating factor α(MFα) signal sequence, downstream the alcoholoxidase 1 ( AOX 1 ) promoter of the pPIC9 vector. The reconstructed vector was subsequently integrated into themethanol yeast P. pastoris strain GS115. The highly expressing strain was selected and the protein sequences andtheir biological activities were assayed. Results The amino acid sequence of rhPF4 was the same as that of naturalone. The rhPF4 can neutralize the anticoagulation effect of heparin in a dose-dependent manner. Conclusion The amount of rhPF4 expressed in methanol yeast meets well the scale for manufacturing. The biological activities of therhPF4 were the same as that of natural PF4.