Dual nuclease and helicase activities of Helicobacter pylori AddAB are required for DNA repair, recombination, and mouse infectivity.

Publication Type:

Journal Article

Source:

The Journal of biological chemistry, Volume 284, Issue 25, p.16759-66 (2009)

Keywords:

2009, Animals, Anti-Bacterial Agents, Basic Sciences Division, Center-Authored Paper, Ciprofloxacin, Comparative Medicine Core Facility, DNA Helicases, DNA repair, DNA, Bacterial, Escherichia coli, Exodeoxyribonucleases, Female, Genomics Core Facility, Helicobacter pylori, Human Biology Division, Humans, MICE, Mice, Inbred C57BL, Mutagenesis, Site-Directed, Protein Structure, Tertiary, Recombinant Proteins, Recombination, Genetic, Scientific Imaging Core Facility, Shared Resources, Virulence

Abstract:

Helicobacter pylori infection of the human stomach is associated with disease-causing inflammation that elicits DNA damage in both bacterial and host cells. Bacteria must repair their DNA to persist. The H. pylori AddAB helicase-exonuclease is required for DNA repair and efficient stomach colonization. To dissect the role of each activity in DNA repair and infectivity, we altered the AddA and AddB nuclease (NUC) domains and the AddA helicase (HEL) domain by site-directed mutagenesis. Extracts of Escherichia coli expressing H. pylori addA(NUC)B or addAB(NUC) mutants unwound DNA but had approximately half of the exonuclease activity of wild-type AddAB; the addA(NUC)B(NUC) double mutant lacked detectable nuclease activity but retained helicase activity. Extracts with AddA(HEL)B lacked detectable helicase and nuclease activity. H. pylori with the single nuclease domain mutations were somewhat less sensitive to the DNA-damaging agent ciprofloxacin than the corresponding deletion mutant, suggesting that residual nuclease activity promotes limited DNA repair. The addA(NUC) and addA(HEL) mutants colonized the stomach less efficiently than the wild type; addB(NUC) showed partial attenuation. E. coli DeltarecBCD expressing H. pylori addAB was recombination-deficient unless H. pylori recA was also expressed, suggesting a species-specific interaction between AddAB and RecA and also that H. pylori AddAB participates in both DNA repair and recombination. These results support a role for both the AddAB nuclease and helicase in DNA repair and promoting infectivity.