酿酒酵母rav2基因的克隆及其在大肠杆菌中的表达  

Cloning and Expression of rav2 Gene from Saccharomyces cerevisiae in Escherichia coli

作　　者：张光明[1] 张震宇[1] 

机构地区：[1]江南大学生物工程学院工业生物技术教育部重点实验室,无锡214122

出　　处：《生物技术通报》2011年第3期191-195,共5页Biotechnology Bulletin

基　　金：国家自然科学基金项目（30970058）

摘　　要：RAVE （regulator of the H ^＋ -ATPase of the vacuolar and endosomal membranes）复合物由Rav1p、Rav2p和Skp1p3（DE3）中进行表达。通过IPTG诱导，SDS-PAGE分析重组菌在诱导后表达出目的蛋白。目的蛋白经Ni-NTA凝胶纯化后再经质谱进一步验证确定为酿酒酵母的Rav2p蛋白。目前国际上还没有有关Rav2p的结构和性质以及RAVE亚基之间相互关系的研究。重组质粒pETDuet-R2的成功构建以及Rav2p的可溶性表达为研究RAVE亚基之间的相互作用以及V-ATP酶的活性调节机理打下基础。The RAVE （ regulator of the H ^＋ -ATPase of the vacuolar and endosomal membranes） complex contains three subunits, Ravlp, Rav2p, and Skplp. The ray2 gene was cloned from Saccharomyces cerevisiae and inserted into corresponding sites of vector pETDuet-1, resulting in the expression plasmid pETDuet-R2. Escherichia coli BL21 （DE3） cells harboring pETDuet-R2 ex- pressed Rav2p after inducted with IPTG. The target protein was analyzed with SDS-PAGE and confirmed by the assay of LC-MS after purified by Ni-NTA resin. There is no report about structure and character of Rav2p so far. This study provided an important basis for investigating the relationship of the three subunits of RAVE and the regulation mechanism of the V-ATPase.

关 键 词：大肠杆菌 克隆表达 RAVE Rav2p 

分 类 号：Q78[生物学—分子生物学]