摘要
将编码正常人肺表面活性物质相关蛋白A1基因的cDNA克隆至酿酒酵母的分泌表达载体pVT10 2U α中 ,构建了重组质粒pVT10 2U α SP A1,转化酵母宿主菌S 78,通过改变培养基的pH值水平 ,经摇瓶培养 ,SDS PAGE结果显示 ,培养上清中SP A1表达量达 40 0mg L以上 ,表达产物分子量为 6 2kD和 32kD。分别以二聚体和单体形式出现。ELISA和Westernblot实验表明表达产物能被抗体特异性识别。生物学活性检测证实其具有调理肺巨噬细胞吞噬E .coli的功能。
The cDNA encoding pulmonary surfactant-associated protein A1 (SP-A1) derived from healthy adult's lung was cloned into the pVT102U/α,expression vector of \%Saccharomyces cerevisiae,\%which contains the yeast α-factor signal sequence,leading to the secretion of expressed protein,and then transformed into \%Saccharomyces cerevisiae\% S-78 (leu2,ura3,rep4) by electroporation.After 2～3 days culture in adequate pH,the expressed SP-A1 accumulated up to 400mg/L in supernatant.The pure proteins were obtained by Sephadex G-25,G-75,Sepharose 4B.The expressed recombinant products,62kD and 32kD,reacted to specific antibody using ELISA and Western blot.The SP-A1 protein expressed in \%Saccharomyces cerevisiae was efficient in enhancing the phagocytosis of \%E.coli\% J5 by alveolar macrophages.
出处
《生物工程学报》
CAS
CSCD
北大核心
2001年第4期410-413,共4页
Chinese Journal of Biotechnology
基金
国家自然科学基金! (3 9670 770 )
广东省自然科学基金!(95 0 5 5 8)
国家863计划项目资助&&
