粪肠球菌群体感应系统luxS基因的研究  

Function analysis of luxS gene relating to quorum sensing system in Enterococcus faecalis

作　　者：邵长林[1,2] 孙忠科[2] 尚伟[1,2] 袁静[2] 廖祥儒[1] 

机构地区：[1]江南大学工业生物技术教育部重点实验室,江苏无锡214122 [2]军事医学科学院疾病预防控制所,北京10007

出　　处：《军事医学》2011年第11期842-846,共5页Military Medical Sciences

基　　金：国家科技重大专项资助项目(2009ZX10004-205);国家自然科学基金资助项目(81071321)

摘　　要：目的研究粪肠球菌群体感应系统luxS基因的功能并体外合成AI-2信号分子,为研究粪肠球菌LuxS/AI-2依赖的群体感应系统提供依据。方法以哈氏弧菌BB170作为报告菌株,检测3株粪肠球菌培养上清的AI-2活性,然后从粪肠球菌中扩增出luxS和pfs基因,测序和比对氨基酸序列,并将两基因插入到pGEX-4T-1表达载体中,转化至大肠杆菌BL21(DE3)中实现表达与纯化,进而以S-腺苷高半胱氨酸为底物、纯化的蛋白或构建的大肠杆菌菌体蛋白为反应酶,体外合成AI-2。结果 3株粪肠球菌均能够分泌有活性的AI-2信号分子;LuxS蛋白序列分析发现其与哈氏弧菌、沙门菌、大肠杆菌具有高度同源性;SAH均可以与纯化的蛋白或构建的大肠杆菌菌体蛋白合成出有活性的AI-2分子。结论粪肠球菌存在LuxS/AI-2依赖的群体感应系统,AI-2的产生依赖于luxS基因。Objective To study the function of luxS gene relating to quorum sensing system and synthesize AI-2 signaling molecules in vitro and to further explore the LuxS/AI-2 dependent quorum sensing system in Enterococcus faecalis.Methods First,Vibrio harveyi BB170 was used as a reporter strain to detect the AI-2 activity of culture supernatants from E.faecalis.Secondly,luxS and pfs genes were amplified from E.faecalis and compared with the amino acid sequences.Then,the two genes were inserted into pGEX-4T-1 expression vector,transformed into Escherichia coli BL21（DE3） for expression and purification.Finally,AI-2 was synthesized in vitro with S-adenosylhomocysteine（SAH） as a substrate and the purified proteins or cell proteins from E.coli as catalytic enzymes for the reaction.Results Three strains of E.faecalis were all able to secrete the active AI-2 signaling molecules.A high LuxS protein homology was analyzed among E.faecalis,V.harveyi,Salmonella typhimurium and E.coli.The active AI-2 molecules could be synthesized by SAH with purified proteins or cell proteins from E.coli.Conclusion LuxS/AI-2 dependent quorum sensing system is found in E.faecalis and the luxS gene is required for AI-2 production.

关 键 词：粪肠球菌 群体感应 LUXS基因 AI-2活性 

分 类 号：R378.2[医药卫生—病原生物学]