结核分枝杆菌rv0394c基因在耻垢分枝杆菌中的表达及其粘附特性的鉴定  

Identification of adhesion characteristic and expression of Mycobacterium tuberculosis rv0394c gene in Mycobacterium smegmatis

作　　者：李华芳[1] 曹俊[1] 刘松艳[1] 陈利苹[1] 刘思国[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/动物细菌病研究室,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2015年第3期199-202,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家自然科学基金(31201920;31272538);兽医生物技术国家重点实验室自主研究(SKLVBP201417);上海市结核病(肺)重点实验室开放基金(2013K03)

摘　　要：本实验室前期研究鉴定出结核分枝杆菌的rv0394c基因产物具有透明质酸酶的活性,但对于该酶在分枝杆菌致病中的作用尚不清楚。为了鉴定该蛋白对细胞是否具有粘附特性,本研究利用大肠杆菌表达纯化的重组蛋白RV0394c与人肺泡上皮细胞(A549细胞)互作,通过激光共聚焦显微镜观察到蛋白对细胞的特异性粘附。为进一步验证该试验结果,将rv0394c基因克隆至大肠杆菌-分枝杆菌穿梭表达质粒p MV262中,将重组质粒电转化于非致病性的耻垢分枝杆菌中进行表达。构建的重组菌与A549细胞进行互作,结果表明重组菌可以粘附于A549细胞表面,而且这种粘附能够被RV0394c重组蛋白所阻断,这些结果表明RV0394c蛋白具有粘附细胞的能力,属于该菌的粘附蛋白之一。RV0394c蛋白粘附特性的鉴定为进一步研究该蛋白在致病性中的作用提供了实验依据。In our previous studies, the RV0394c protein of Mycobacterium tuberculosis was identified to have the activity of hyaluronidase, but the function of the protein is unknown. In order to identify the adhesion characteristic of the protein, RV0394e recombinant protein expressed fiom E.coli were interacted with human lung epithelial （A549） cell and proved to possess the adhesion ability. To further confirm the result, the rv0394c gene was cloned into E.coli-mycobacterimn shuttle vector pMV262, and the resultant recombinant plasmid was transformed into Mycobacterium smegmatis by electroporation for expression, and the result showed that the recombinant bacteria was able to adhere to A549 cell surface when recombinant Mycobacterimn was incubated with A549 cell. Furthermore, the adhesion was also able to be blocked by RV0394c protein. These data demonstrat that the RV0394c protein is one of adhesion protein of Mycobacterium tuberculosis. The identification of RV0394c protein adhesion characteristic provides a basis for further study the role of the protein in pathogenicity of the bateria.

关 键 词：结核分枝杆菌 rv0394c基因 pMV262 耻垢分枝杆菌 粘附 

分 类 号：S852.61[农业科学—基础兽医学]