Varela Lorena, Bell Christian H, Armitage Judith P, Redfield Christina
Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK.
Biomol NMR Assign. 2016 Oct;10(2):373-8. doi: 10.1007/s12104-016-9703-x. Epub 2016 Jul 29.
Rhodobacter sphaeroides has emerged as a model system for studies of the complex chemotaxis pathways that are a hallmark of many non-enteric bacteria. The genome of R. sphaeroides encodes two sets of flagellar genes, fla1 and fla2, that are controlled by three different operons. Each operon encodes homologues of most of the proteins required for the well-studied E. coli chemotaxis pathway. R. sphaeroides has six homologues of the response regulator CheY that are localized to and are regulated by different clusters of chemosensory proteins in the cell and have different effects on chemotaxis. CheY6 is the major CheY stopping the fla1 flagellar motor and associated with a cytoplasmically localised chemosensory pathway. CheY3 and CheY4 are associated with a membrane localised polar chemosensory cluster, and can bind to but not stop the motor. CheY6 and either CheY3 or CheY4 are required for chemotaxis. We are using NMR spectroscopy to characterise and compare the structure and dynamics of CheY3 and CheY6 in solution. We are interested in defining the conformational changes that occur upon activation of these two proteins and to identify differences in their properties that can explain the different functions they play in chemotaxis in R. sphaeroides. Here we present the (1)H, (13)C and (15)N assignments for CheY3 in its active, inactive and Mg(2+)-free apo form. These assignments provide the starting point for detailed investigations of the structure and function of CheY3.
球形红细菌已成为研究复杂趋化途径的模型系统，这些途径是许多非肠道细菌的一个标志。球形红细菌的基因组编码两组鞭毛基因，fla1和fla2，它们由三个不同的操纵子控制。每个操纵子编码了大肠杆菌趋化途径中大多数所需蛋白质的同源物。球形红细菌有六个反应调节蛋白CheY的同源物，它们定位于细胞内不同的化学感受蛋白簇并受其调节，并且对趋化作用有不同影响。CheY6是使fla1鞭毛马达停止转动的主要CheY，与细胞质定位的化学感受途径相关。CheY3和CheY4与膜定位的极性化学感受簇相关，并且可以结合但不能使马达停止转动。趋化作用需要CheY6以及CheY3或CheY4中的一种。我们正在使用核磁共振光谱来表征和比较溶液中CheY3和CheY6的结构和动力学。我们感兴趣的是确定这两种蛋白质激活时发生的构象变化，并识别它们性质上的差异，这些差异可以解释它们在球形红细菌趋化作用中发挥的不同功能。在这里，我们展示了处于活性、非活性和无Mg(2+)的脱辅基形式的CheY3的(1)H、(13)C和(15)N归属。这些归属为详细研究CheY3的结构和功能提供了起点。
