前脂蛋白乙二酰转移酶基因(lgt)与霍乱弧菌噬菌体VP3的受体相关  

Prolipoprotein diacylglyceryl transferase (lgt) is a gene associated with receptor of V.cholerae phage VP3

作　　者：王多春[1] 郭晓琳[2] 刁保卫[1] 梁未丽[1] 邱海燕[1] 高守一[1] 阚飙[1] 

机构地区：[1]中国疾病预防控制中心传染病预防控制所卫生部医学分子细菌学重点实验室 [2]兰州医学院附属二院

出　　处：《中华微生物学和免疫学杂志》2004年第7期564-567,共4页Chinese Journal of Microbiology and Immunology

基　　金："973"国家重大基础研究项目 (G19990 5 410 2 ) ;国家自然科学基金项目 (3 0 2 71169)

摘　　要：目的　研究霍乱弧菌噬菌体VP3的受体相关基因 ,确定前脂蛋白乙二酰转移酶基因在霍乱菌株可被VP3感染中的作用。方法　采用转座子 (TnphoA)插入 ,筛选对VP3感染产生抗性的突变株。Southernblot鉴定突变株染色体中TnphoA的拷贝数 ,利用反向PCR扩增TnphoA插入位点旁侧研究菌株的染色体序列。对扩增产物测序确定插入位点基因。另外 ,克隆被破坏基因的完整序列 ,转入该突变株 ,以反式互补实验确定回补突变株对VP3的敏感性。结果　获得 1株VP3的抗性转座突变株 ,Southernblot确定TnphoA在其中为单拷贝插入 ,反向PCR(IPCR)扩增得到插入位点处的结构基因为霍乱弧菌lgt基因 ,其表达产物为前脂蛋白乙二酰转移酶 (Lgt)。反式互补表明突变株恢复了对VP3的敏感性。结论　前脂蛋白乙二酰转移酶在赋予霍乱菌株噬菌体VP3的敏感性方面有重要作用 ,可能为VP3转染的受体合成中的必需酶类。Objective To study the receptor associated genes of V.cholerae phage VP3,and determine the role of prolipoprotein diacylglyceryl transferase in the process of VP3 infects to V.cholerae. Methods Transponson insertion was used to screen mutants resistant to VP3. Southern blot was used to identify the insertion copy number of transponson,inverse PCR was used to amplify the flanking sequence of insertion site located on the strain chromosome. The amplified production was sequenced and determined the gene of insertion site. Meanwhile,clone the integrated gene which was disrupted,complementation experiment was used to identify the sensibility of mutant to VP3. Results A VP3-resistant mutant was obtained and the insertion was confirmed with a single insertion by Southern blot,using inverse PCR,the structural gene of insert site was amplified as lgt in V.cholerae coding prolipoprotein diacylglyceryl transferase. Complementation experiment confirmed the role of lgt in VP3 infection,it′s VP3 susceptibility was fully restored. Conclusion Prolipoprotein diacylglyceryl transferase plays an important role in the susceptibility of VP3, it suggests an essential enzyme in the receptor synthesis.

关 键 词：前脂蛋白 乙二酰转移酶基因 lgt 噬菌体VP3 受体 霍乱弧菌 染色体 

分 类 号：R346[医药卫生—基础医学]