作　　者： ; ; ; ; ; ;

机构地区： 广西大学生命科学与技术学院

出　　处： 《广西科学》 2008年第2期184-188,共5页

摘　　要： 以酿酒酵母(Saccharomyces cerevisiae)基因组DNA为模板,通过PCR扩增得到蔗糖酶基因(suc2),并将其克隆到表达载体pSE380中,将得到的重组质粒pSE-suc2转化进E.coli BL21中,利用镍金属螯合层析方法分析测定其酶学性质。重组菌株的SDS-PAGE结果显示重组蔗糖酶基因(suc 2)有60kDa目的蛋白出现,纯化的SDS-PAGE分析得到均一的蛋白条带;重组蔗糖酶的Km值为47.73mmol/L,最大反应速率为79.59mg还原糖mg-1蛋白min-1,最适温度为42℃,最适pH值为5.5,Zn2+、Cu2+对重组蔗糖酶酶活有较强的抑制作用,Ba2+、Mg2+、Mn2+对重组蔗糖酶酶活稍有激活作用。 The gene encoded invertase from Saccharomyces cerevisiae was amplified by PCR, and inserted into the expression vector pSE380, then introduced into E. coli BL21. The recombinant protein was purified by immobilized metal affinity chromatography. It was presented as a single protein band on SDS-PAGE with molecular weight of 60kD. The recombinant enzyme followed typical Michaelis-Menten kinetics with an apparent Km of 47. 73mmol/L. Its Vmax value was 79. 59mg reducing sugar mg^-1protein min^-1. The optimum activity of the recombinant invertase was found to be at pH value of Ba^2＋.Mg^2＋ ,Mn^2＋, while 5.5 and 42℃C. The invertase activity was increased slightly by addition inhibited by Zn^2＋ .Cu^2＋.

关 键 词： 蔗糖酶 克隆 表达 酶学性质

领　　域： [生物学]