Shrivastava Rashmi, Das Dibya Ranjan, Wiker Harald G, Das Amit Kumar
Department of Biotechnology, Indian Institute of Technology, Kharagpur 721302, India.
Biochem Biophys Res Commun. 2006 Jun 16;344(4):1327-33. doi: 10.1016/j.bbrc.2006.04.019. Epub 2006 May 2.
Two-component systems (TCSs) are the major signalling pathway in bacteria and represent potential drug targets. Among the 11 paired TCS proteins present in Mycobacterium tuberculosis H37Rv, the histidine kinases (HKs) Rv0600c (HK1) and Rv0601c (HK2) are annotated to phosphorylate one response regulator (RR) Rv0602c (TcrA). We wanted to establish the sequence-structure-function relationship to elucidate the mechanism of phosphotransfer using in silico methods. Sequence alignments and codon usage analysis showed that the two domains encoded by a single gene in homologous HKs have been separated into individual open-reading frames in M. tuberculosis. This is the first example where two incomplete HKs are involved in phosphorylating a single RR. The model shows that HK2 is a unique histidine phosphotransfer (HPt)-mono-domain protein, not found as lone protein in other bacteria. The secondary structure of HKs was confirmed using "far-UV" circular dichroism study of purified proteins. We propose that HK1 phosphorylates HK2 at the conserved H131 and the phosphoryl group is then transferred to D73 of TcrA.
双组分系统（TCSs）是细菌中的主要信号传导途径，也是潜在的药物靶点。在结核分枝杆菌H37Rv中存在的11对TCS蛋白中，组氨酸激酶（HKs）Rv0600c（HK1）和Rv0601c（HK2）被注释为可磷酸化一个应答调节因子（RR）Rv0602c（TcrA）。我们希望建立序列-结构-功能关系，以利用计算机方法阐明磷酸转移机制。序列比对和密码子使用分析表明，同源HKs中由单个基因编码的两个结构域在结核分枝杆菌中已被分离为单独的开放阅读框。这是两个不完整的HKs参与磷酸化单个RR的首个例子。该模型表明，HK2是一种独特的组氨酸磷酸转移（HPt）单结构域蛋白，在其他细菌中未发现其单独存在。通过对纯化蛋白进行“远紫外”圆二色性研究，证实了HKs的二级结构。我们提出，HK1在保守的H131位点磷酸化HK2，然后磷酸基团转移至TcrA的D73位点。
