长双歧杆菌NCC2705中胆盐水解酶基因的克隆表达及其特性的研究  

Cloning of bile salt hydrolase in Bifidobacterium longum NCC2705 and description of its characteristics

作　　者：李云梅[1,2] 尚伟[2,3] 郭晶[1,2] 邹大阳[2] 杜双奎[1] 袁静[2] 

机构地区：[1]西北农林科技大学食品科学与工程学院,陕西杨凌712100 [2]军事医学科学院疾病预防控制所,北京100071 [3]江南大学工业生物技术教育部重点实验室,江苏无锡214122

出　　处：《军事医学》2012年第8期586-589,594,共5页Military Medical Sciences

基　　金：国家“十二五”科技重大专项资助项目(2011ZX10004-001)

摘　　要：目的克隆并过表达胆盐水解酶基因,研究胆盐水解酶对菌体胆盐耐受性和胆固醇降解率的影响等特性。方法采用常规原核表达方法在大肠杆菌BL21(DE3)和长双歧杆菌NCC2705中过表达胆盐水解酶基因,并进行长双歧杆菌NCC2705和pMgap-bsh/NCC2705的胆盐耐受性和降胆固醇实验。结果在大肠杆菌中成功表达了NCC2705的胆盐水解酶基因,经纯化的GST空标签和GST融合蛋白的SDS-PAGE电泳结果显示,胆盐水解酶的相对分子质量约为35×103;构建的过表达bsh的菌株pMgap-bsh/NCC2705比标准菌株NCC2705表现出了略高的胆盐耐受性和胆固醇降解率。结论所构建的高效表达胆盐水解酶菌株可能有利于提高菌体的胆盐耐受性和降胆固醇能力,益生菌的降胆固醇途径不是单一的,培养基中是否含有胆盐对胆固醇的降解率影响较大。Objective To clone and overexpress bile salt hydrolase of Bifidobacterium longum NCC2705 in E. coli BL21 （ DE3 ） and to test the bile salt tolerance and cholesterol-lowering efficiency of B. longum NCC2705 and pMgap bile salt hydrolase gene （bsh）/NCC2705. Methods A conventional prokaryotic expression method was used to clone and express bile salt hydrolase gene in E. coli BL21 （DE3） and B. longum NCC2705. Bile salt tolerance and cholesterol-lower- ing ability of B. longum NCC2705 and pMgap-bsh/NCC2705 were explored. Results The bile salt hydrolase gene was suc- cessfully expressed,and GST-bsh fusion protein was well purified. The relative molecular mass of bile salt hydrolase was about 35 x lO3 according to the results of SDS-PAGE gel electrophoresis of purified GST-bsh fusion protein and GST empty tag of expression broth supernatant of pGEX-4T-1-bsh/BL21 （ DE3 ）. Bile salt tolerance and cholesterol-lowering ability of pMgap-bsh/NCC2705 was higher than that of the standard strain NCC2705. Conclusion The strains of overexpressed bile salt bydrolase may help to improve the bile tolerance and cholesterol-lowering ability of strains. Probioticis does not lower cholesterol in a single way, which is related to the bile salt concentration of the medium.

关 键 词：长双歧杆菌NCC2705 胆盐水解酶 克隆表达 特性 

分 类 号：R378.2[医药卫生—病原生物学]