From the Center for Protein Science and Crystallography, School of Life Sciences, Faculty of Science, Chinese University of Hong Kong, Shatin, Hong Kong.
Shenzhen Research Institute, The Chinese University of Hong Kong, Shenzhen 518057, China, and.
J Biol Chem. 2018 Sep 7;293(36):13961-13973. doi: 10.1074/jbc.RA118.002263. Epub 2018 Jul 10.
Bacterial flagella are rotary nanomachines that contribute to bacterial fitness in many settings, including host colonization. The flagellar motor relies on the multiprotein flagellar motor-switch complex to govern flagellum formation and rotational direction. Different bacteria exhibit great diversity in their flagellar motors. One such variation is exemplified by the motor-switch apparatus of the gastric pathogen , which carries an extra switch protein, FliY, along with the more typical FliG, FliM, and FliN proteins. All switch proteins are needed for normal flagellation and motility in , but the molecular mechanism of their assembly is unknown. To fill this gap, we examined the interactions among these proteins. We found that the C-terminal SpoA domain of FliY (FliY) is critical to flagellation and forms heterodimeric complexes with the FliN and FliM SpoA domains, which are β-sheet domains of type III secretion system proteins. Surprisingly, unlike in other flagellar switch systems, neither FliY nor FliN self-associated. The crystal structure of the FliY-FliN complex revealed a saddle-shaped structure homologous to the FliN-FliN dimer of , consistent with a FliY-FliN heterodimer forming the functional unit. Analysis of the FliY-FliN interface indicated that oppositely charged residues specific to each protein drive heterodimer formation. Moreover, both FliY-FliM and FliY-FliN associated with the flagellar regulatory protein FliH, explaining their important roles in flagellation. We conclude that uses a FliY-FliN heterodimer instead of a homodimer and creates a switch complex with SpoA domains derived from three distinct proteins.
细菌鞭毛是旋转纳米机器，在许多环境中有助于细菌适应，包括宿主定殖。鞭毛马达依赖于多蛋白鞭毛马达开关复合物来控制鞭毛的形成和旋转方向。不同的细菌在其鞭毛马达方面表现出极大的多样性。这种变化的一个例子是胃病原体的马达开关装置，它携带一个额外的开关蛋白 FliY，以及更典型的 FliG、FliM 和 FliN 蛋白。所有的开关蛋白对于正常的鞭毛形成和运动都是必需的，但它们的组装分子机制尚不清楚。为了填补这一空白，我们研究了这些蛋白质之间的相互作用。我们发现 FliY（FliY）的 C 端 SpoA 结构域对于鞭毛形成至关重要，并与 FliN 和 FliM SpoA 结构域形成异二聚体复合物，这些结构域是 III 型分泌系统蛋白的 β-折叠结构域。令人惊讶的是，与其他鞭毛开关系统不同，FliY 和 FliN 都不自我关联。FliY-FliN 复合物的晶体结构揭示了一种鞍形结构，与 的 FliN-FliN 二聚体同源，这与 FliY-FliN 异二聚体形成功能单元一致。对 FliY-FliN 界面的分析表明，每个蛋白特有的相反电荷残基驱动异二聚体的形成。此外，FliY-FliM 和 FliY-FliN 都与鞭毛调节蛋白 FliH 相关联，这解释了它们在鞭毛形成中的重要作用。我们得出结论， 使用 FliY-FliN 异二聚体代替同源二聚体，并利用三个不同蛋白的 SpoA 结构域创建开关复合物。
