Abstract
Neisseria meningitidis (Nm) is a Gram-negative oral commensal that opportunistically can cause septicaemia and/or meningitis. Here, we overexpressed, purified and characterized the Nm DNA repair/recombination helicase RecG (RecGNm) and examined its role during genotoxic stress. RecGNm possessed ATP-dependent DNA binding and unwinding activities in vitro on a variety of DNA model substrates including a Holliday junction (HJ). Database searching of the Nm genomes identified 49 single nucleotide polymorphisms (SNPs) in the recGNm including 37 non-synonymous SNPs (nsSNPs), and 7 of the nsSNPs were located in the codons for conserved active site residues of RecGNm. A transient reduction in transformation of DNA was observed in the Nm ΔrecG strain as compared to the wildtype. The gene encoding recGNm also contained an unusually high number of the DNA uptake sequence (DUS) that facilitate transformation in neisserial species. The differentially abundant protein profiles of the Nm wildtype and ΔrecG strains suggest that expression of RecGNm might be linked to expression of other proteins involved in DNA repair, recombination and replication, pilus biogenesis, glycan biosynthesis and ribosomal activity. This might explain the growth defect that was observed in the Nm ΔrecG null mutant.
Highlights
Neisseria meningitidis (Nm), or the meningococcus, is a Gram-negative bacterium that frequently colonizes the human oropharynx
To test the specific unwinding activity on branched DNA substrates, increasing concentrations of RecGNm were incubated with end-labelled DNA substrates in the presence of 2 mM ATP and Mg2+
RecG is a double-stranded DNA translocase and helicase thought to play multiple roles in cellular processes including initiation of origin-dependent DNA replication, remodelling, regressing and restarting replication forks stalled at DNA lesions [22,58]
Summary
Neisseria meningitidis (Nm), or the meningococcus, is a Gram-negative bacterium that frequently colonizes the human oropharynx. In individuals who lack bactericidal antibodies, Nm can enter the bloodstream, cross the blood-brain barrier, and cause septicaemia and/or meningitis [1]. We are interested in how Nm cells survive on the oral mucosal surface, in the bloodstream and at the meninges, where it is exposed to reactive oxygen and nitrogen species that are typically highly genotoxic [2]. It has been reported that Neisseria may be less proficient in DNA base excision repair (BER) than E. coli and lacks an SOS response to DNA damage [2,3] These features, and its genetic tractability due to its constitutive competence for transformation and short generation time, make Nm
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