Shiomi Daisuke, Zhulin Igor B, Homma Michio, Kawagishi Ikuro
Division of Biological Science, Graduate School of Science, Nagoya University, Chikusa-ku, Japan.
J Biol Chem. 2002 Nov 1;277(44):42325-33. doi: 10.1074/jbc.M202001200. Epub 2002 Jul 5.
Adaptation to persisting stimulation is required for highly sensitive detection of temporal changes of stimuli, and often involves covalent modification of receptors. Therefore, it is of vital importance to understand how a receptor and its cognate modifying enzyme(s) modulate each other through specific protein-protein interactions. In the chemotaxis of Escherichia coli, adaptation requires methylation of chemoreceptors (e.g. Tar) catalyzed by the CheR methyltransferase. CheR binds to the C-terminal NWETF sequence of a chemoreceptor that is distinct from the methylation sites. However, little is known about how CheR recognizes its methylation sites or how it is distributed in a cell. In this study, we used comparative genomics to demonstrate that the CheR chemotaxis methyltransferase contains three structurally and functionally distinct modules: (i) the catalytic domain common to a methyltransferase superfamily; (ii) the N-terminal domain; and (iii) the beta-subdomain of the catalytic domain, both of which are found exclusively in chemotaxis methyltransferases. The only evolutionary conserved motif specific to CheR is the positively charged face of helix alpha2 in the N-terminal domain. The disulfide cross-linking analysis suggested that this face interacts with the methylation helix of Tar. We also demonstrated that CheR localizes to receptor clusters at cell poles via interaction of the beta-subdomain with the NWETF sequence. Thus, the two chemotaxis-specific modules of CheR interact with distinct regions of the chemoreceptor for targeting to the receptor cluster and for recognition of the substrate sites, respectively.
为了对刺激的时间变化进行高灵敏度检测,需要适应持续的刺激,这通常涉及受体的共价修饰。因此,了解受体及其同源修饰酶如何通过特定的蛋白质-蛋白质相互作用相互调节至关重要。在大肠杆菌的趋化作用中,适应需要由CheR甲基转移酶催化的化学感受器(如Tar)的甲基化。CheR与化学感受器的C末端NWETF序列结合,该序列与甲基化位点不同。然而,关于CheR如何识别其甲基化位点或其在细胞中的分布方式知之甚少。在这项研究中,我们使用比较基因组学来证明CheR趋化作用甲基转移酶包含三个结构和功能不同的模块:(i)甲基转移酶超家族共有的催化结构域;(ii)N末端结构域;(iii)催化结构域的β亚结构域,这两个结构域仅在趋化作用甲基转移酶中发现。CheR唯一进化保守的基序是N末端结构域中α2螺旋的带正电荷表面。二硫键交联分析表明,该表面与Tar的甲基化螺旋相互作用。我们还证明,CheR通过β亚结构域与NWETF序列的相互作用定位于细胞极的受体簇。因此,CheR的两个趋化作用特异性模块分别与化学感受器的不同区域相互作用,以靶向受体簇并识别底物位点。