中温α-淀粉酶基因在枯草芽孢杆菌中的高效表达及其发酵条件优化  

High level expression of medium α-amylase in Bacillus subtilis and optimization of fermentation conditions

作　　者：张士彬[1,2] 陈景奇[2] 崔艳艳[1] 谢能中[3] 王敏[1] 张大伟[2] 

机构地区：[1]天津科技大学生物工程学院,天津300457 [2]中国科学院天津工业生物技术研究所,天津300308 [3]广西科学院,非粮生物质酶解国家重点实验室,国家非粮生物质能源工程技术研究中心,广西南宁530007

出　　处：《食品与发酵工业》2016年第4期50-56,共7页Food and Fermentation Industries

基　　金：国家自然科学基金(No.31370089,21446008,31400079);广西省科技支撑项目(14125008-2-22)

摘　　要：以枯草芽孢杆菌为表达系统,以中温α-淀粉酶为目标蛋白,研究了启动子和信号肽对外源蛋白表达分泌的影响。分别选取了4种启动子以及8种信号肽来进行筛选,结果表明,启动子PApr E的强度最高,信号肽SPnpr E的分泌效率最好。针对重组菌株1A751Q31进行系统的发酵优化,在72 h发酵液中α-淀粉酶酶活达到380U/m L。在7.5 L发酵罐中进行放大实验,发酵液中α-淀粉酶酶活最高达到853 U/m L。以上研究表明,α-淀粉酶适合在枯草芽孢杆菌中进行表达,同时表明枯草芽孢杆菌是良好的异源蛋白表达系统。In this study,the effect of promoter and signal peptide on the expression of heterologous model protein,α-amylase,was studied by using Bacillus subtilis as the expression system. Four promoters and eight signal peptides were compared,and the best promoter and signal peptide for the expression and secretion of α-amylase were identified. The results indicated that PApr Eand SPnpr Ewere the most suitable for the production of α-amylase in B. subtilis. The fermentation of the recombinant B. subtilis 1A751Q31 was then optimized,and the activity of α-amylase in the supernate achieved 380 U / mL. Fed-batch fermentation in a 7. 5 L fermentor was carried out,and the activity ofα-amylase in the supernate reached 853 U / mL. All the studies in this paper indicated that it was suitable for the expression of α-amylase using B. subtilis as the expression system,and B. subtilis was a suitable system for expression of heterologous proteins.

关 键 词：枯草芽孢杆菌 Α-淀粉酶 启动子 信号肽 发酵优化 

分 类 号：TQ925[轻工技术与工程—发酵工程]