FlgY, PflA, and PflB form a spoke-ring network in the high-torque flagellar motor of <i>Helicobacter pylori</i>.

Abstract

<i>Helicobacter pylori</i> has evolved distinct flagellar motility to colonize the human stomach. Rotation of the <i>H. pylori</i> flagella is driven by one of the largest known bacterial flagellar motors. In addition to the core motor components found in <i>Escherichia coli</i> and <i>Salmonella enterica</i>, the flagellar motor in <i>H. pylori</i> possesses many accessories that enable the bacteria to penetrate the gastric mucus layer. Here, we utilize cryoelectron tomography with molecular genetics and biochemical approaches to characterize three accessory proteins, FlgY, PflA, and PflB, and their roles in <i>H. pylori</i> flagellar assembly and motility. Comparative analyses of in situ flagellar motor structures from <i>pflA</i>, <i>pflB</i>, and <i>flgY</i> mutants and wild-type <i>H. pylori</i> reveal that FlgY forms a 13-fold proximal spoke-ring around the MS-ring and that PflA and PflB form an 18-fold distal spoke-ring enclosing 18 torque-generating stator complexes. We build a pseudoatomic model of the <i>H. pylori</i> motor by leveraging AlphaFold-predicted structures, protein-protein interactions, and <i>in situ</i> motor structures. Our model suggests that the FlgY spoke-ring functions as a bearing around the rotating MS-ring and as a template for stabilizing the PflA-PflB spoke-ring, thus enabling the recruitment of 18 stator complexes for high-torque generation. Overall, our study sheds light on how this spoke-ring network between the MS-ring and stator complexes enables the unique motility of <i>H. pylori</i>. As these accessory proteins are conserved in the phylum Campylobacterota, our findings apply broadly to a better understanding of how polar flagella help bacteria thrive in gastric and enteric niches.