构建相应ompC基因质粒了解孔道蛋白突变与大肠埃希菌耐药的关系  

Identification and molecular analysis of relationship between mutations in ompC and E.coli antibiotic resistance

作　　者：戎建荣[1] 王淑峰[1] 李连青[1] Alison S.low Ian R.Booth 

机构地区：[1]山西省临床检验中心,太原030012 [2]英国苏格兰阿伯丁大学细胞与分子生物研究院

出　　处：《中国卫生检验杂志》2010年第5期985-987,1064,共4页Chinese Journal of Health Laboratory Technology

基　　金：山西省医学留学人员重点基金项目(20000940)

摘　　要：目的:研究和分析细胞膜孔道蛋白OmpC的突变与大肠埃希菌对抗生素生耐药的关系。方法:将野生株E.coliK-12的低拷贝孔道蛋白ompC基因转入两株临床分离的对抗生素耐药性不同的大肠埃希菌菌体细胞中(1号和7号),检测比较它们在转化前后对不同抗生素的MIC值。同时克隆1号和7号菌株的孔道蛋白ompC基因,构建相应的质粒分别转化入HN705受体细胞内,观察其细菌对药物的最低抑菌浓度(MIC)值的变化,了解孔道蛋白改变与细菌耐药的相关性。外膜蛋白采用SDS-PAGE电泳和质谱仪进行分析。结果:临床分离的1号大肠埃希菌在获得野生株ompC基因前后其MIC值无显著变化;临床分离菌株7号大肠埃希菌在获得野生株ompC蛋白基因后,对卡巴培能、美罗培南和亚胺培南的MIC值显著下降,对药物的敏感性增强,说明野生株ompC基因在7号菌体内表达以后恢复了细胞膜的部分通透功能,使更多的抗生素进入细菌体内达到作用靶位,降低了细菌对抗生素的耐药水平。结论:大肠埃希菌孔道蛋白ompC氨基酸序列的改变影响相应蛋白的表达,使细胞膜通透性降低,增加细菌对抗生素的耐药性。Objective：To identify and investigate if the variant OmpC being expressed by the clinical strains was playing a role in their antibiotic resistance.Methods：A low copy number plasmid containing the wild type E.coli K-12 ompC gene was transformed into the two different clinical isolates.Increased expression of OmpC,indicating expression of the chromosomal copy and the plasmid copy of the gene,was confirmed by carrying out outer membrane preparations and running SDS-PAGE gels.Results：To In the isolate 1 there were no significant changes in MIC with or without the wild type OmpC The isolate 7 however did show significant decreases in MICs for the carbapenem antibiotics,meropenem and imipenem,when wild type OmpC was expressed.It appears that the expression of wild type OmpC in the last isolate at least partially restores permeability allowing antibiotics into the cell and reducing the level of antibiotic resistance.This result indicates that at least one of the amino acid changes present in the last isolate 7.Conclusion：Prliminary data suggest that changes in the amino acid sequence of the contribute to antibiotic resistance via the modulation of ompC active.

关 键 词：细胞膜孔道蛋白 克隆 转化 质粒构建 突变 

分 类 号：R378.9[医药卫生—病原生物学]