Abstract
BackgroundHelicobacter pylori HP0958 protein (FlgZ) prevents the rapid turnover of RpoN (σ54), a transcription factor required for expression of several flagellar genes in H. pylori. FlgZ possesses a zinc-ribbon domain (DUF164) that contains two conserved CXXC motifs which coordinate a zinc ion and is thought to interact with nucleic acids or proteins. Two conserved cysteine residues in FlgZ (Cys-202 and Cys-223) were replaced with serine to assess their significance in FlgZ function. After confirming the importance of the CXXC motifs in the DUF164 domain of FlgZ, the distribution of DUF164 proteins and RpoN homologs in other bacteria was examined to determine if a correlation existed for the concurrence of the two proteins.ResultsLevels of RpoN were greatly reduced in H. pylori strains that expressed the FlgZC202S or FlgZC223S variants. The FlgZC202S variant, but not the FlgZC223S variant, accumulated at levels similar to the wild-type protein. DUF164 proteins are not universally distributed and appear to be absent in several major bacterial taxa, including Cyanobacteria as well as Alpha-, Beta- and Gammaproteobacteria. With the exception of the Actinobacteria, members of which generally lack RpoN, genes encoding DUF164 proteins and RpoN are frequently found in the same genome. Interestingly, many of the DUF164 proteins in Actinobacteria and Bacteroidetes lack most or even all of the conserved cysteine residues.ConclusionsThese findings suggest the importance of the zinc-ribbon domain of FlgZ in protecting RpoN from turnover. Since many bacteria that possess a DUF164 protein also contain RpoN, DUF164 proteins may have roles in RpoN protection or function in other bacteria.
Highlights
Helicobacter pylori HP0958 protein (FlgZ) prevents the rapid turnover of RpoN (s54), a transcription factor required for expression of several flagellar genes in H. pylori
FlgZ are important for function and stability To determine if the conserved cysteine residues in the zinc-ribbon domain of FlgZ are critical for the function of the protein, mutations were introduced in each of the conserved CXXC motifs and the phenotypes of strains expressing the resulting FlgZ variants were examined
Cys-202 and Cys-223 were changed to serine residues and the resulting FlgZ variants were expressed in a mutant strain of H. pylori 43504 in which flgZ was disrupted with a kanamycin-resistance cassette
Summary
Helicobacter pylori HP0958 protein (FlgZ) prevents the rapid turnover of RpoN (s54), a transcription factor required for expression of several flagellar genes in H. pylori. FlgZ possesses a zinc-ribbon domain (DUF164) that contains two conserved CXXC motifs which coordinate a zinc ion and is thought to interact with nucleic acids or proteins. Many of the DUF164 proteins in Actinobacteria and Bacteroidetes lack most or even all of the conserved cysteine residues. Transcriptional activation of the RpoN-dependent flagellar genes in H. pylori requires the response regulator FlgR and its cognate histidine kinase FlgS [12,14]. Transcription of the H. pylori RpoN regulon requires the putative RpoN chaperone and zinc-ribbon domain protein HP0958 [19,20], which we hereafter refer to as FlgZ (flagellar-associated zinc-ribbon domain protein)
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