芝田硫化叶菌新型α-淀粉酶基因工程菌表达条件的优化  

Studies on the Expression Condition of Engineered Strain of the Novel α-Amylase from Surfolbus shibatae in E.coli.

作　　者：刘莉[1] 吴襟[1] 陈炜[1] 张树政[1] 

机构地区：[1]中国科学院微生物研究所酶学研究室,北京100080

出　　处：《中国生物工程杂志》2003年第1期70-74,共5页China Biotechnology

基　　金：北京市自然科学基金项目 (No .5 982 0 10 );微生物资源国家重点实验室 (No .9810 2 )课题

摘　　要：对本室构建的生产海藻糖的基因工程菌株 ,即来自古细菌芝田硫化叶菌B1 2的新型α 淀粉酶基因的工程菌的表达外源蛋白条件进行宿主菌 ,培养基 ,诱导时间 ,诱导温度和诱导菌浓的起始OD60 0 等条件的优化 ,发现在宿主菌为大肠杆菌DH5α ,培养基为改进的LB ,诱导时间 4小时 ,诱导温度 41℃ ,OD60 0 为 0 6～ 0 8时 ,基因工程菌ESBAM中新型α 淀粉酶的表达量最高 ,新型α 淀粉酶活力也最高 ,与重组酶MTSase一起作用底物直链淀粉 ,经HLPC检测在反应产物中有海藻糖的生成。Expression conditions of engineering bacteria E.coli DH5α/SBAM expressed novel α |amylase from Sulfolobus shibatae B12 were optimized.Search for how expression conditions to affect engineering bacterial expressing level of protein.We modified various conditions which affect expression,developed a success technique for the ESBAM to express target protein at higher level.The optimum culture conditions were the host strain DH5α,medium LB ++,induction temperature 41℃,induction time 4 hours and the initial concentration OD 600 0 6-0 8 of the engineering strain ESBAM respectively.Trehalose was detected by HPLC after amylose was acted by the two recombinant enzymes cooperatively,which were the novel α |amylase and the maltooligosyltrehalose synthase(MTSase) from Sulfobobus shibatae. ;

关 键 词：芝田硫化叶菌 新型α-淀粉酶 基因工程菌 表达条件 优化 酶活合成 古细菌 海藻糖 

分 类 号：TQ929[轻工技术与工程—发酵工程] Q933[生物学—微生物学]