Roles of linker region flanked by transmembrane and peptidoglycan binding region of PomB in energy conversion of theVibrioflagellar motor

Abstract

ABSTRACTThe energy converting complex of the sodium-driven flagellar motor in bacteria comprises two proteins, PomA and PomB, whose transmembrane regions form ion conducting channels and is called the stator complex. The transmembrane protein PomB is attached to the cell wall by its periplasmic region and has a plug segment following the transmembrane helix to prevent ion flux. PomB (Δ41-120), which lacks the periplasmic region from E41 to K120 immediately following its transmembrane region shows similar motility as that of wild-type PomB. In this study, three deletion mutants after the plug region, PomB (Δ61-120), PomB (Δ61-140), and PomB (Δ71-150), were generated and those deletion mutants were examined for their functionality. PomB (Δ61-120) conferred similar motility as that of the wild-type protein, whereas the other two mutants showed almost no motility in soft agar plate; however, we observed some swimming cells with speed similar to that of the wild-type cells. To observe dominance of wild-type proteins, we introduced the two PomB mutants into a wild-type strain, and its ability to swim was not affected by the mutants. Then, we purified the mutant PomAB complexes to confirm the stator formation. When we introduced the PomB mutations in the plug region, the reduced motility by the deletion was rescued, suggesting that the stator was activated. Our results indicate that the deletion prevents stator from transformation to an active form; however, the linker and plug regions from E41 to S150 are not essential for the motor function of PomB but are important for its regulation.IMPORTANCEThe stator complex of flagella consists of PomA and PomB proteins and interacts with the rotor complex. PomB has a peptidoglycan binding (PGB) domain to fix the stator for generation of torque. PomB is attached to the cell wall only when the stator is activated by interaction between the cytoplasmic region of PomA and the rotor protein FliG. The flexible linker of PomB, which is a naturally unfolded region, is flanked by the peptidoglycan-binding (PGB) domain and transmembrane region. The plug region, which interacts with the periplasmic loops of PomA to prevent activation of the stator, is located next to its transmembrane region. In this study, we reveal the role of the flexible linker in activation of the stator complex.