酿酒酵母ATP合酶δ亚基融合蛋白的构建、表达及生物学功能分析  

Construction,Expression and Functional Analysis of Haemagglutinin-tagged δ Subunit of Saccharomyces cerevisiae Mitochondrial ATP Synthase

作　　者：于立权[1] 闫智慧[2] 蒋伶活[1,2] 

机构地区：[1]天津大学药物科学与技术学院,天津300072 [2]天津医科大学天津市基础医学研究中心,天津300070

出　　处：《生物技术通报》2009年第8期99-103,共5页Biotechnology Bulletin

基　　金：国家自然科学基金项目(30870107)

摘　　要：利用分子克隆手段构建了酿酒酵母ATP合酶δ亚基和流感病毒血凝素(haemagglutinin,HA)标签融合蛋白的表达质粒。该融合蛋白在酿酒酵母细胞中能够正常表达,而且能够互补编码δ亚基的ATP16基因缺失株在利用非发酵性碳源方面的表型缺陷,表明该融合蛋白具有野生型δ亚基的功能。同时,构建了在大肠杆菌细胞中表达该δ亚基的ScAtp16p-His6融合蛋白,并用纯化的融合蛋白在家兔中制备了其多抗血清。结果表明此多抗可以很好地与ScAtp16p-His6和HA-ScAtp16p两种融合蛋白特异性结合。这些研究材料的获得为深入研究ATP合酶的解偶联机制和磷酸化调控机理奠定了基础。A construct expressing the HA-tagged δ subunit of F1F0 ATP synthase was constructed. Functional complementation study showed that the fusion protein could complement the function of the wild-type δ subunit in the utilization of non-fermentable carbon sources in yeast cells. Meanwhile, antiserum against inclusion bodies containing bacterial expressed ScATP16-His6 proteins was generated, and this polyclonal antibody could recognize fusion proteins ScATP16-His6 and HA-ScAtp16p. These studies provide a basis to further study the uncoupling mechanism between F1 and F0 portions of ATP syuthase and its regulation by protein phosphorylation in the mitochondrion.

关 键 词：酿酒酵母F1F0-ATP合酶δ亚基 融合蛋白 功能分析 

分 类 号：Q78[生物学—分子生物学]