Characterization of the periplasmic region of PomB, a Na+-driven flagellar stator protein in Vibrio alginolyticus |
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Authors: | Li Na Kojima Seiji Homma Michio |
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Affiliation: | Division of Microbiology, Graduate School of Life Science, Northwest A&F University, Yanglin, Japan. z47616a@cc.nagoya-u.ac.jp |
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Abstract: | The stator proteins PomA and PomB form a complex that couples Na+ influx to torque generation in the polar flagellar motor of Vibrio alginolyticus. This stator complex is anchored to an appropriate place around the rotor through a putative peptidoglycan-binding (PGB) domain in the periplasmic region of PomB (PomBC). To investigate the function of PomBC, a series of N-terminally-truncated and in-frame mutants with deletions between the transmembrane (TM) segment and the PGB domain of PomB was constructed. A PomBC fragment consisting of residues 135 to 315 (PomBC5) formed a stable homodimer and significantly inhibited the motility of wild-type cells when overexpressed in the periplasm. A fragment with an in-frame deletion (PomBΔL) of up to 80 residues retained function, and its overexpression with PomA impaired cell growth. This inhibitory effect was suppressed by a mutation at the functionally critical Asp (D24N) in the TM segment of PomB, suggesting that a high level of Na+ influx through the mutant stator causes the growth impairment. The overproduction of functional PomA/PomBΔL stators also reduced the motile fractions of the cells. That effect could be slightly relieved by a mutation (L168P) in the putative N-terminal α-helix that connects to the PGB domain without affecting the growth inhibition, suggesting that a conformational change of the region including the PGB domain affects stator assembly. Our results reveal common features of the periplasmic region of PomB/MotB and demonstrate that a flexible linker that contains a “plug” segment is important for the control of Na+ influx through the stator complex as well as for stator assembly. |
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