title: The GTPase activity of FlhF is dispensable for flagellar localization, but not motility, in Pseudomonas aeruginosa.
abstract: The opportunistic human pathogen Pseudomonas aeruginosa uses two surface organelles, flagella and pili, for motility and adhesion in biotic and abiotic environments. Polar flagellar placement and number are influenced by FlhF, which is a signal recognition particle (SRP)-type GTPase. The FlhF proteins of Bacillus subtilis and Campylobacter jejuni were recently shown to have GTPase activity. However, the phenotypes associated with flhF deletion and/or mutation differ between these organisms and P. aeruginosa, making it difficult to generalize a role for FlhF in pseudomonads. In this study, we confirmed that FlhF of P. aeruginosa binds and hydrolyzes GTP. We mutated FlhF residues that we predicted would alter nucleotide binding and hydrolysis and determined the effects of these mutations on FlhF enzymatic activity, protein dimerization, and bacterial motility. Both hydrolytically active and inactive FlhF point mutants restored polar flagellar assembly, as seen for wild-type FlhF. However, differential effects on flagellar function were observed in single-cell assays of swimming motility and flagellar rotation. These findings indicate that FlhF function is influenced by its nucleotide binding and hydrolytic activities and demonstrate that FlhF affects P. aeruginosa flagellar function as well as assembly.
authors.affiliation: Departments of Medicine, Yale University School of Medicine, New Haven, Connecticut, USA.
authors.forename: Maren
authors.initials: M
authors.lastname: Schniederberend
authors.forename: Kholis
authors.initials: K
authors.lastname: Abdurachim
authors.forename: Thomas Scott
authors.initials: TS
authors.lastname: Murray
authors.forename: Barbara I
authors.initials: BI
authors.lastname: Kazmierczak
mesh_terms: D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D010861:Fimbriae, Bacterial / Q000235:genetics / Q000502:physiology; D005407:Flagella / Q000235:genetics / Q000502:physiology*; D020558:GTP Phosphohydrolases / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D020559:Monomeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D009068:Movement; D017354:Point Mutation; D055503:Protein Multimerization; D011550:Pseudomonas aeruginosa / Q000201:enzymology / Q000235:genetics / Q000502:physiology*; D017384:Sequence Deletion; D015398:Signal Transduction
chemical_list: D001426:Bacterial Proteins; C078317:flhF protein, Bacteria; D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases; D020559:Monomeric GTP-Binding Proteins