Title: Directed evolution of the stators of the flagellar motor - exploring phenamil resistance.

Abstract:
We used error-prone PCR to introduce single point mutations into the sodium-powered Vibrio alginolyticus / Escherichia coli chimeric stator PotB and selected for motors that exhibited motility in the presence of the sodium-channel inhibitor phenamil. We found single mutations that enable motility under phenamil occurred at two sites: 1) the transmembrane domain of PotB, corresponding to the TM region of the PomB stator from V. alginolyticus, and 2) near the peptidoglycan (PG) binding region that corresponds to the C-terminal region of the MotB stator from E. coli. Using phylogenetic logistic regression, we found correlation between natural residue variation and ion source at positions corresponding to PotB F22Y, but not at other sites. Our results demonstrate that it is not only the pore region of the stator that moderates motility in the presence of ion-channel blockers.