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Abstract
The pathogen Helicobacter pylori has evolved distinct flagellar motility to colonize the human stomach of approximately half the world’s population. Assembly and rotation of the H. pylori flagella are driven by one of the largest known bacterial flagellar motors. In addition to the core motor components in Escherichia coli and Salmonella enterica, the flagellar motor in H. pylori possesses many accessories that presumably facilitate the high torque needed to penetrate the gastric mucus layer. H. pylori hp0838 encodes a previously uncharacterized lipoprotein and is in an operon with flgP, which encodes a motor accessory protein. Deletion analysis of hp0838 in H. pylori B128 showed that the gene is not required for motility in soft agar medium, but the mutant displayed a reduced growth rate and an increased sensitivity to bacitracin. A variant of the Δhp0838 mutant containing a frameshift mutation in pflA, which resulted in paralyzed flagella, displayed wild-type growth rate and resistance to bacitracin, suggesting the fitness defect and antibiotic sensitivity of the Δhp0838 mutant are dependent on flagellar rotation. Comparative analysis of in-situ structures of the wild type and Δhp0838 mutant motors revealed the Δhp0838 mutant motor lacked a previously undescribed ring structure with 18-fold symmetry located near the outer membrane, which we named the FapH (flagellar accessory protein in Helicobacter pylori) ring. ΔfapH Δfur and ΔfapH ΔlpxF mutants were found to have less sensitivity to bacitracin, suggesting mechanisms responsible for suppressing the sensitivity phenotype. Flagellar motor accessory proteins FlgY, PflA, and PflB, and their roles in H. pylori flagellar assembly and motility are characterized. Comparative analyses of pflA, pflB, and flgY mutants and wild-type H. pylori B128 reveal that FlgY forms a 13-fold proximal spoke-ring around the MS-ring, while PflA and PflB form an 18-fold distal spoke-ring at the periphery of the motor. Our model suggests that the FlgY 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 torque-generating stator complexes in each motor. As these accessory proteins are conserved in H. pylori and other members of the phylum Campylobacterota, our findings apply broadly to an understanding of how polar flagella help bacteria thrive in distinct niches.