Institutes of Biomedical Science, Fudan University, Shanghai 200433, China.
Metallomics. 2013 Oct;5(10):1448-57. doi: 10.1039/c3mt00163f.
TroR is a putative metal-dependent regulatory protein that has been linked to the virulence of the human pathogen Treponema pallidum. It shares high homology with the well-known iron-dependent regulatory protein DtxR from Corynebacterium diphtheriae, as well as the manganese-dependent MntR from Bacillus subtilis. However, it has been uncertain whether manganese or zinc is the natural cofactor of TroR to date. Herein, we established an efficient method named "double-fusion tagging" to obtain soluble TroR for the first time. A series of studies, including ICP, CD, fluorescence, ITC, and electrophoresis mobility shift assay (EMSA), were performed to resolve the discrepancies in its metal-binding specificity. In addition, bioinformatic analysis as well as mutation studies were carried out to find the genetic relationships of TroR with its homology proteins. In conclusion, our findings indicate that TroR is a manganese-dependent rather than a zinc-dependent regulatory protein.
TroR 是一种假定的金属依赖性调节蛋白，与人类病原体苍白密螺旋体的毒力有关。它与众所周知的来自白喉棒状杆菌的铁依赖性调节蛋白 DtxR 以及来自枯草芽孢杆菌的锰依赖性 MntR 具有高度同源性。然而，迄今为止，尚不确定锰还是锌是 TroR 的天然辅因子。在此，我们首次建立了一种名为“双融合标记”的有效方法来获得可溶性 TroR。进行了一系列研究，包括 ICP、CD、荧光、ITC 和电泳迁移率变动分析（EMSA），以解决其金属结合特异性中的差异。此外，还进行了生物信息学分析和突变研究，以发现 TroR 与其同源蛋白的遗传关系。总之，我们的研究结果表明 TroR 是一种锰依赖性而不是锌依赖性的调节蛋白。
