大肠埃希菌外膜蛋白T水解人防御素-4作用  

Hydrolytic effect of Escherichia coli outer membrane protein T on human-beta-defensin-4

作　　者：屈娅荣[1] 赵铁[1] 曹曦[1] 尤帅[1] 余晶仪[1] 温扬明[1] 张立科[1] 曹虹[1] 

机构地区：[1]南方医科大学公共卫生与热带医学学院微生物系,广东广州510515

出　　处：《中国公共卫生》2013年第5期696-698,共3页Chinese Journal of Public Health

基　　金：国家自然科学基金(30972637);南方医科大学公卫学院院长基金(GW201101)

摘　　要：目的探讨外膜蛋白T(OmpT)在大肠埃希菌CFT073致尿路感染中的作用机制。方法以人膀胱癌上皮细胞5637的cDNA为模版,扩增人防御素-4(HBD-4)基因,连接至质粒pET-28a,重组质粒经转化、诱导和纯化处理后获得His-HBD-4融合蛋白;采用琼脂糖扩散法和最小抑菌浓度方法检测HBD-4对ompT野生株、敲除株和回补株的抗菌活性,并分别观察比较ompT野生株CFT073、敲除株和回补株水解HBD-4的差异。结果通过测序鉴定及双酶切电泳证实成功构建HBD-4原核表达载体pET-28a-HBD-4,获得HBD-4体外表达菌株;HBD-4对CFT073与ompT敲除株的最小抑菌浓度不同,野生株为10μg/mL,敲除株为6μg/mL;与HBD-4共同孵育后,敲除株的生长状态受到抑制,但对野生株的影响较小。结论初步证实ompT作为毒力因子参与对人固有免疫系统成分的水解,在大肠杆菌致尿路感染的过程中发挥重要作用。Objective To examine the role of Escherichia coli outer membrane protein T in urinary tract infection （UTI）. Methods The gene encoding human-beta-defensin-4 （HBD-4） was amplifide by PCR and then cloned into vec- tor pET-28a to construct pET-28a/HBIM. The His-HBD4 expression was induced by isopropyl-[3-D-thioglactopyranoside （IPTG）. The antimicrobial activity of the HBD-4 was tested by minimal inhibitory concentration（MIC） and agarose dif- fusion method was used to observe the differences in hydrolysis of HBD-4 induced by CFT073 and CFT073 AornpT. Re- suits The recombinant expression vetor pET-28a/HBD4 was successfully constructed. There were differences in hydrol- ysis of HBD-4 induced by wild-type, the complementation and mutant strains. The MIC of HBD4 for wild-type was 10 p^g/ml but that of for mutant strain was 6 p,g/ml. After co-incubated with HBD-4, the growth of mutant strain was inhibited but that of the wild-type strain was not affected obviously. Conclusion Escherichia coli outer membrane pro- tein Thydrolyzes HBD-4, which plays an important role in the pathogenesis of uropathogenic Escherichia coli-induced UTI.

关 键 词：大肠埃希菌 外膜蛋白T(OmpT) 尿路感染 防御素-4(HBD-4) 

分 类 号：R37[医药卫生—病原生物学]