Characterization of PomA periplasmic loop and sodium ion entering in stator complex of sodium-driven flagellar motor
Characterization of PomA periplasmic loop and sodium ion entering in stator complex of sodium-driven flagellar motor
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PomA 周质环和钠离子进入钠驱动鞭毛电机定子复合体的表征
DOI：
10.1093/jb/mvz102
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发表时间：
2019
期刊：
影响因子：
--
通讯作者：
Homma Michio
中科院分区：
文献类型：
--
作者：
Nishikino Tatsuro;Iwatsuki Hiroto;Mino Taira;Kojima Seiji;Homma Michio
细菌鞭毛马达是由离子流驱动的旋转纳米机器。海洋弧菌鞭毛定子复合体由两种蛋白质PomA和PomB组成，在海洋弧菌中进行能量传递。PomA是一种四跨膜（TM）蛋白，并且TM 2和TM 3之间的胞质区域（环2 -3）与转子蛋白FliG相互作用以产生扭矩。TM 1和TM 2（环1 -2）以及TM 3和TM 4（环3 -4）之间的周质区域是定子的跨膜离子通道的入口处的候选者。在这项研究中，我们纯化的定子复合物与半胱氨酸替代周质环和评估的蛋白质与生物素马来酰亚胺（BM）的反应性。BM容易修饰环3 - 4中的Cys残基，但几乎不标记环1 -2中的Cys残基。我们不能纯化由PomBΔ41- 120和WT-PomA组成的插塞去除定子（ΔL定子），但可以用回路1 - 2的PomA-D31 C或TM的PomB-D24 N来纯化ΔL定子。当离子通道关闭时，PomA和PomB强烈相互作用。当离子通道打开时，PomA与PomB的相互作用不那么紧密。plug和loop 1 - 2区域调节定子的这种激活，这取决于钠离子与PomB的D24残基的结合。
The bacterial flagellar motor is a rotary nanomachine driven by ion flow. The flagellar stator complex, which is composed of two proteins, PomA and PomB, performs energy transduction in marineVibrio. PomA is a four transmembrane (TM) protein and the cytoplasmic region between TM2 and TM3 (loop2–3) interacts with the rotor protein FliG to generate torque. The periplasmic regions between TM1 and TM2 (loop1–2) and TM3 and TM4 (loop3–4) are candidates to be at the entrance to the transmembrane ion channel of the stator. In this study, we purified the stator complex with cysteine replacements in the periplasmic loops and assessed the reactivity of the protein with biotin maleimide (BM). BM easily modified Cys residues in loop3–4but hardly labelled Cys residues in loop1–2. We could not purify the plug deletion stator (ΔL stator) composed of PomBΔ41–120and WT-PomA but could do the ΔL stator with PomA-D31C of loop1–2or with PomB-D24N of TM. When the ion channel is closed, PomA and PomB interact strongly. When the ion channel opens, PomA interacts less tightly with PomB. The plug and loop1–2region regulate this activation of the stator, which depends on the binding of sodium ion to the D24 residue of PomB.