1株古细菌海藻糖合成酶系在枯草芽孢杆菌中的诱导表达  

Heterologous Expression of Enzymes for Trehalose Synthesis from Sulfolobus acidocaldarius in Bacillus subtilis

作　　者：王珊瑛[1,2] 李由然[1,2] 顾正华[1,2] 张梁[1,2] 丁重阳[1,2] 石贵阳[1,2] 

机构地区：[1]江南大学粮食发酵工艺与技术国家工程实验室,江苏无锡214122 [2]江南大学生物工程学院,江苏无锡214122

出　　处：《食品科学》2016年第21期124-130,共7页Food Science

基　　金：国家自然科学基金青年科学基金项目(31401674);江南大学自主重点项目(JUSRP51503)

摘　　要：来源于古细菌嗜酸硫化叶菌(Sulfolobus acidocaldarius)的麦芽寡糖基海藻糖合成酶(maltooligosylterhalose synthase,MTSase)和麦芽寡糖基海藻糖水解酶(maltooligosyltrehalose trehalohydrolase,MTHase)联合作用,可以利用淀粉为底物生成海藻糖。本研究构建了6种枯草芽孢杆菌一大肠杆菌穿梭表达载体,将密码子优化后的MTSase和MTHase编码基因在木糖异构酶基因的启动子及其阻遏蛋白介导下实现了在枯草芽孢杆菌中的功能表达,木糖启动子分别来源于枯草芽孢杆菌(Bacillus subtilis)、巨大芽孢杆菌(Bacillus megaterium)、地衣芽孢杆菌(Bacillus licheniformis)。在以5 g/L甘油为碳源,24 g/L蛋白胨为氮源时,菌体培养8 h后加入终质量浓度为6 g/L的木糖,37℃诱导16 h后重组MTSase、MTHase产生联合作用,海藻糖转化率达到33.57%。实现了MTSase、MTHase在枯草芽孢杆菌中的功能表达。Trehalose has great potential applications in both the medical and food industries due to its unique properties. Enzymatic transformation is superior to extraction from yeast cells for the industrial production of trehalose. Starch can be transformed into trehalose via maltooligosylterhalose synthase（MTSase） and maltooligosyltrehalose trehalohydrolase（MTHase） from Sulfolobus acidocaldarius. In this experiment, we constructed six recombinant expression plasmids harboring the codon-optimized MTSase and MTHase encoding genes from Sulfolobus acidocaldarius, which could be expressed under the control of the xylose operons from Bacillus subtilis, Bacillus megaterium and Bacillus licheniformis. The recombinant plasmids were transformed into Bacillus subtilis, respectively. The effects of different culture conditions on the production of trehalose was investigated and the optimal culture conditions were determined as follows： glycerol concentration as the best carbon source, 5 g/L; peptone concentration as the best nitrogen source, 24 g/L; xylose concentration, 6 g/L; incubation duration, 8 h; and induction temperature, 37 ℃; induction time, 16 h. Experiments carried out under these conditions gave a conversion rate of trehalose of 33.57%. This study has successfully achieved the functional expression of MTSase and MTHase in Bacillus subtilis.

关 键 词：穿梭质粒 嗜酸硫化叶菌 枯草芽孢杆菌 诱导表达 

分 类 号：Q814[生物学—生物工程]