凝结芽孢杆菌中β-半乳糖苷酶基因的克隆及表达  

Molecular cloning and expression of a novel β-galactosidase from Bacillus coagulans

作　　者：郑兆娟[1] 徐颖[1] 石磊[1] 欧阳嘉[1] 

机构地区：[1]南京林业大学化学工程学院江苏省生物质绿色燃料与化学品重点实验室,江苏南京210037

出　　处：《南京林业大学学报（自然科学版）》2015年第6期35-39,共5页Journal of Nanjing Forestry University：Natural Sciences Edition

基　　金：国家自然科学基金青年基金项目(31300487;31200443);江苏省自然科学基金青年项目(BK20130970);江苏高校优势学科建设工程资助项目(PAPD)

摘　　要：从1株快速水解乳糖发酵产酸的凝结芽孢杆菌NL01的基因组上克隆获得了1个新的β-半乳糖苷酶基因,该基因长度为1 998 bp,其编码的氨基酸序列与已经报道的β-半乳糖苷酶相似度低于40%。将该β-半乳糖苷酶基因整合到表达载体pETDuet-1上,并在大肠杆菌BL21(DE3)中进行重组表达,β-半乳糖苷酶粗酶酶活为1190μmol/(min·mg),经镍柱纯化获得的重组β-半乳糖苷酶酶活为6664μmol/(min·mg)。利用该重组β-半乳糖苷酶水解乳糖,经TLC和HPLC分析显示该酶具有将乳糖水解为葡萄糖和半乳糖的活力,是一种新型的β-半乳糖苷酶。A novel β-galactosidase gene was cloned fromBacillus coagulansNL01 which had ability to hydrolyze lactoseinto glucose and galactose in this study. The length of the β-galactosidase gene was 1 998 bp, and its coding sequenceshowed very low identity with other reported β-galactosidase. The gene was cloned into pETDuet-1 and expressed inEscherichia coliBL21（DE3）. The crude enzyme activity was 119.0 μmol/（min·mg）, and the purified enzyme activitywas 666.4 μmol/（min·mg） after Ni-NTA purification. It was indicated from the analysis results of the hydrolysis prod-uct obtained by the purified β-galactosidase that this novel β-galactosidase presented high activity toward lactose con-version into glucose and galactose.

关 键 词：凝结芽孢杆菌 Β-半乳糖苷酶 表达纯化 乳糖水解 

分 类 号：Q815[生物学—生物工程] TQ93[化学工程]