The gene for the Salmonella FliJ protein is flanked by those of the FliI ATPase and the hook length control protein FliK, as part of the FliE operon. Flagellar genes in H. pylori are not contained in such large

operons, but are scattered throughout the genome [23, 32]. HP0256 is flanked by an adenylosuccinate synthetase gene (purA/HP0255) as well as two outer membrane protein genes (omp7/HP0252 and omp8/HP0254), and three hypothetical genes, one of which encodes a predicted secreted protein (HP0257) and the other a predicted Bromosporine datasheet integral membrane protein (HP0258). When comparing HP0256 with homologues from related species, it did not appear that any one domain of the protein was more or less conserved (Figure 2). This agrees with previous studies of FliJ data suggesting that the entire protein is necessary for function [28]. As this bioinformatic analysis suggested HP0256 could be a FliJ homologue, we generated a HP0256 mutant by inserting a chloramphenicol resistance marker into the gene by allelic exchange as described in Methods. Growth rates and plate morphology of the HP0256 mutant were indistinguishable from the wild-type (data not www.selleckchem.com/products/cb-839.html shown). Ablation of the HP0256 gene reduces motility Motility plate assay indicated that the HP0256 mutant was significantly less motile than the wild-type

(Figure 3). A similar phenotype was consistently observed in two H. pylori wild-type strains and their derivative HP0256 mutants (Figure 3), indicating that the reduced motility was not a strain-specific effect. However, the mutants retained some motility. In Salmonella, lack of FliJ abolishes motility [27], suggesting that HP0256 may not be a FliJ homologue as AG-120 cell line initially hypothesized. Complementation of a Salmonella FliJ mutant was attempted by introduction of the HP0256 gene expressed from an E. coli vector promoter. Motility plate selleck compound assay

indicated that motility was not restored in the Salmonella fliJ mutant, indicating that HP0256 was unlikely to be a functional FliJ homologue in Helicobacter pylori (data not shown). We complemented the P79-derivative HP0256 mutant, by expressing the HP0256 gene, integrated into the chromosome, under the control of the flaA promoter (Figure 3). Restoration of motility in the complemented mutant confirmed that the partial loss of motility in the mutant was due only to the lack of the HP0256 gene product. Figure 3 The ablation of the HP0256 gene impairs motility in H. pylori that may be restored by complementation, when hp0256 is put under the control of the promoter of flaA. Motility plate assay were performed four times. A. CCUG17874 wild-type strain; B. CCUG17874-hp0256KO; C. P79 wild-type strain; D. P79-hp0256KO; E. P79-hp0256KO complemented with pIR0601; F. P79-hp0256KO with empty vector (control).