连接肽对融合蛋白ELP[I]_(30)-linker-eGFP相变的影响  

The Impact of Linker Peptide on Phase Transition of Fusion Protein ELP[I]_(30)-linker-eGFP

作　　者：张立超[1] 李军明[1] 葛高顺[1] 孔令岭[1] 胡学军[1] 

机构地区：[1]大连大学医学院,中国辽宁大连116622

出　　处：《生命科学研究》2013年第5期401-405,共5页Life Science Research

基　　金：国家自然科学基金资助项目(31070822);辽宁省教育厅高校科研计划项目(L2010015);大连大学博士启动基金资助项目(2012-3)

摘　　要：研究G4S和Poly N连接肽对融合蛋白ELP[I]30-linker-eGFP相变的影响.将编码两种不同连接肽G4S和Poly N的绿色荧光蛋白(enhanced green fluorescent protein,eGFP)基因克隆到pET28-ELP[I]30表达载体中,在宿主菌E.coli BLR(DE3)中经IPTG诱导表达ELP[I]30-linker-eGFP,通过可逆相变循环(inverse transition cycling,ITC)及镍柱亲和层析纯化ELP[I]30-linker-eGFP蛋白.结果显示,成功构建、表达具有活性的两种连接肽的融合蛋白ELP[I]30-linker-eGFP,连接肽G4S使融合蛋白产生不可逆相变,而Poly N不影响融合蛋白可逆相变,该研究对类弹性蛋白标签的应用具有指导意义.To investigate the effection of different linker peptides on the fusion ELP[Ⅰ]30-eGFP proteins＇ bioac- tivity and phase transition, the eGFP genes with encoding G4S and Poly N linker were inserted into pET28- ELP[Ⅰ]30expression vector respectively. The fusion proteins were expressed in E. coli BLR（DE3） induced by IPTG respectively, and were confirmed via SDS-PAGE and Western blot. The fusion proteins were purified via ITC （Inverse transition cycling） and Nickel column affinity chromatography respectively. The fusion pro- teins ELP[Ⅰ]30-1inker-eGFP were successfully expressed and purified with activity. The linker peptide G4S could cause fusion protein an irreversible phase change, and the linker peptide Poly N didn＇t affect the fusion pro- tein reversible phase transition, which laid the foundation for the application of ELP[Ⅰ]30.

关 键 词：连接肽 融合蛋白 纯化标签 类弹性蛋白 绿色荧光蛋白 

分 类 号：Q819[生物学—生物工程]