Bao Yi-ge, Qi Zi-fang, Bao Lang
School of Clinical Medicine, School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2009 Dec;29(12):2371-4.
To clone and express the Rv3871 gene related to the virulent protein secretion of Mycobacterium tuberculosis and analyze its molecular structure, function and homology using bioinformatic approach.
A pair of primers was designed to amplify the Rv3871 gene, which was subcloned into the prokaryotic plasmid pET32a(+). The recombinant plasmid was identified by sequence analysis and the expressed recombinant protein by SDS-PAGE. The structure, function and homology alignment of Rv3871 were analyzed comparatively against other mycobacteria.
The restriction fragments through molecular cloning matched perfectly in size with our prediction. The gene sequence was consistent with the corresponding sequence in GenBank. The expression protein was detected by SDS-PAGE with a molecular weight of 84 kD. Two FtsK/SpoE III domains were found by bioinformatic analysis. The homology results showed distinct differences between Rv3871 of the pathogenic M. tuberculosis and its counterparts in non-pathogenic mycobacteria.
Molecular cloning, expression and sequencing identify the structural and functional characteristics of Rv3871. The structural and functional differences of the gene between pathogenic and non-pathogenic mycobacteria identified by bioinformatics provide some evidence for the pathogenesis and drug targets of tuberculosis.
克隆并表达与结核分枝杆菌毒力蛋白分泌相关的Rv3871基因，采用生物信息学方法分析其分子结构、功能及同源性。
设计一对引物扩增Rv3871基因，将其亚克隆至原核质粒pET32a(+)。通过序列分析鉴定重组质粒，用SDS-PAGE检测表达的重组蛋白。与其他分枝杆菌比较分析Rv3871的结构、功能及同源性比对情况。
通过分子克隆获得的限制性片段大小与预测完全匹配。基因序列与GenBank中的相应序列一致。SDS-PAGE检测到表达蛋白分子量为84 kD。生物信息学分析发现两个FtsK/SpoE III结构域。同源性结果显示致病性结核分枝杆菌的Rv3871与其在非致病性分枝杆菌中的对应物存在明显差异。
分子克隆、表达及测序确定了Rv3871的结构和功能特征。生物信息学鉴定出致病性和非致病性分枝杆菌该基因在结构和功能上的差异，为结核病的发病机制和药物靶点提供了一些证据。
