Department of Microbiology, Monash University, Clayton, Victoria, 3800, Australia.
Department of Physics and Optical Science, University of North Carolina at Charlotte, Charlotte, NC, USA.
Sci Rep. 2017 Oct 18;7(1):13435. doi: 10.1038/s41598-017-13421-0.
Rotation of the bacterial flagellum is powered by a proton influx through the peptidoglycan (PG)-tethered stator ring MotA/B. MotA and MotB form an inner-membrane complex that does not conduct protons and does not bind to PG until it is inserted into the flagellar motor. The opening of the proton channel involves association of the plug helices in the periplasmic region of the MotB dimer into a parallel coiled coil. Here, we have characterised the structure of a soluble variant of full-length Helicobacter pylori MotB in which the plug helix was engineered to be locked in a parallel coiled coil state, mimicking the open state of the stator. Fluorescence resonance energy transfer measurements, combined with PG-binding assays and fitting of the crystal structures of MotB fragments to the small angle X-ray scattering (SAXS) data revealed that the protein's C-terminal domain has a PG-binding-competent conformation. Molecular modelling against the SAXS data suggested that the linker in H. pylori MotB forms a subdomain between the plug and the C-terminal domain, that 'clamps' the coiled coil of the plug, thus stabilising the activated form of the protein. Based on these results, we present a pseudo-atomic model structure of full-length MotB in its activated form.
细菌鞭毛的旋转是由质子通过肽聚糖（PG）连接的定子环 MotA/B 流入驱动的。MotA 和 MotB 形成一个内膜复合物，在插入鞭毛马达之前，它不传导质子，也不与 PG 结合。质子通道的打开涉及到 MotB 二聚体在周质区域的插塞螺旋的平行卷曲螺旋的缔合。在这里，我们对全长幽门螺杆菌 MotB 的可溶性变体的结构进行了表征，其中插塞螺旋被设计为锁定在平行卷曲螺旋状态，模拟定子的打开状态。荧光共振能量转移测量，结合 PG 结合测定和 MotB 片段的晶体结构拟合到小角 X 射线散射（SAXS）数据表明，该蛋白的 C 末端结构域具有 PG 结合能力构象。针对 SAXS 数据的分子建模表明，幽门螺杆菌 MotB 的连接子在插塞和 C 末端结构域之间形成一个亚结构域，“夹住”插塞的卷曲螺旋，从而稳定蛋白的激活形式。基于这些结果，我们提出了全长 MotB 在其激活形式下的拟原子模型结构。
