National Laboratory of Protein Engineering and Plant Genetic Engineering, College of Life Sciences, Peking University, Beijing, China.
Mol Microbiol. 2010 Mar;75(6):1484-94. doi: 10.1111/j.1365-2958.2010.07069.x. Epub 2010 Feb 10.
Sinorhizobium meliloti DctB is a typical transmembrane sensory histidine kinase, which senses C(4)-dicarboxylic acids (DCA) and regulates the expression of DctA, the DCA transporter. We previously reported the crystal structures of its periplasmic sensory domain (DctBp) in apo and succinate-bound states, and these structures showed dramatic conformational changes at dimeric level. Here we show a ligand-induced dimeric switch in solution and a strong correlation between DctBp's dimerization states and the in vivo activities of DctB. Using site-directed mutagenesis, we identify important determinants for signal perception and transduction. Specifically, we show that the ligand-binding pocket is essential for DCA-induced 'on' activity of DctB. Mutations at different sections of DctBp's dimerization interface can lock full-length DctB at either 'on' or 'off' state, independent of ligand binding. Taken together, these results suggest that DctBp's signal perception and transduction occur through a 'ligand-induced dimeric switch', in which the changes in the dimeric conformations upon ligand binding are responsible for the signal transduction in DctB.
苜蓿中华根瘤菌 DctB 是一种典型的跨膜感应组氨酸激酶，能够感应 C(4)-二羧酸（DCA）并调节 DCA 转运蛋白 DctA 的表达。我们之前报道了其周质感应结构域（DctBp）在apo 和琥珀酸结合状态下的晶体结构，这些结构在二聚体水平上表现出显著的构象变化。在这里，我们在溶液中展示了配体诱导的二聚体开关，并发现 DctBp 的二聚化状态与 DctB 的体内活性之间存在很强的相关性。通过定点突变，我们确定了信号感知和转导的重要决定因素。具体来说，我们表明配体结合口袋对于 DCA 诱导的 DctB“开启”活性是必需的。DctBp 二聚化界面不同部位的突变可以使全长 DctB 锁定在“开启”或“关闭”状态，而不依赖于配体结合。总之，这些结果表明，DctBp 的信号感知和转导是通过“配体诱导的二聚体开关”发生的，在这种开关中，配体结合引起的二聚体构象变化负责 DctB 中的信号转导。
