Characterization of the meningococcal DNA glycosylase Fpg involved in base excision repair

Tibballs, Katrina Louise Bø; Ambur, Ole Herman; Alfsnes, Kristian; Homberset, Håvard; Frye, Stephan A.; Davidsen, Tonje; Tønjum, Tone

doi:10.1186/1471-2180-9-7

Cited by 10 publications

(10 citation statements)

References 45 publications

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“…While we found that MutM has glycosylase/lyase activity against 8-oxoG·C (Fig. 2 E ), Nth failed to remove this lesion (Table 1), which is consistent with previous results in E. coli and N. meningitidis (Morland et al, 2002, Tibballs et al, 2009). MutM excised 8-oxoG·C with the highest rate constant observed ( k cl , 0.1 ± 0.006 s −1 ) for this enzyme with any substrate tested.…”

Section: Resultssupporting

confidence: 92%

“…E. coli MutM was initially named Fpg because formamidopyrimidine (FAPY) was its first characterised substrate (Chetsanga & Lindahl, 1979, Boiteux et al , 1989). However, as the E. coli and meningococcal enzymes both recognise other DNA lesions including 8-oxoG (Tibballs et al, 2009, Morland et al , 2002), we refer to this enzyme as MutM (Nghiem et al , 1988). Nothing is known about the activity of the putative meningococcal Nth (encoded by NMB0533), which shares 72% amino acid identity with the E. coli enzyme that recognises a broad range of oxidised pyrimidines (Dizdaroglu et al , 1993).…”

Section: Resultsmentioning

confidence: 99%

“…The other enzyme, NExo, shares 3′-deoxyribose phosphodiesterase (3′-RPase, which processes 3′-Ald) activity with NApe (Carpenter et al , 2007), but has evolved to be a specialised 3′-phosphatase (Silhan et al ., 2011). Additionally, the bacterium is known to possess the bi-functional glycosylase, MutM (also called Fpg), which excises 8-oxoG (Tibballs et al , 2009), and MutY which is an anti-mutator responsible for excising adenine (A) mismatched with 8-oxoG (Davidsen et al , 2005).…”

Section: Introductionmentioning

confidence: 99%

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A network of enzymes involved in repair of oxidative DNA damage in Neisseria meningitidis

Nagorska

Šilhán

et al. 2012

Molecular Microbiology

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SummaryAlthough oxidative stress is a key aspect of innate immunity, little is known about how host-restricted pathogens successfully repair DNA damage. Base excision repair is responsible for correcting nucleobases damaged by oxidative stress, and is essential for bloodstream infection caused by the human pathogen, Neisseria meningitidis. We have characterized meningococcal base excision repair enzymes involved in the recognition and removal of damaged nucleobases, and incision of the DNA backbone. We demonstrate that the bi-functional glycosylase/lyases Nth and MutM share several overlapping activities and functional redundancy. However, MutM and other members of the GO system, which deal with 8-oxoG, a common lesion of oxidative damage, are not required for survival of N. meningitidis under oxidative stress. Instead, the mismatch repair pathway provides back-up for the GO system, while the lyase activity of Nth can substitute for the meningococcal AP endonuclease, NApe. Our genetic and biochemical evidence shows that DNA repair is achieved through a robust network of enzymes that provides a flexible system of DNA repair. This network is likely to reflect successful adaptation to the human nasopharynx, and might provide a paradigm for DNA repair in other prokaryotes.

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Section: Resultssupporting

confidence: 92%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A network of enzymes involved in repair of oxidative DNA damage in Neisseria meningitidis

Nagorska

Šilhán

et al. 2012

Molecular Microbiology

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“…This is especially true for the pathogenic Neisseria strains, where mutators are frequently isolated from epidemic-associated, invasive strains of N. meningitidis compared to related strains that colonize the nasopharynx and rarely cause invasive disease (38). Disrupted genes in N. meningitidis that produce a mutator phenotype include mutS and mutL as well as the fpg and mutY glycosylases of base excision repair and 8-oxo-guanine repair pathways (6,37,38,53). The role of mutators in the natural history of Gc is less well understood, but we presume that mutator clinical isolates of Gc do exist and will be found to lack active MutS or MutL.…”

Section: Discussionmentioning

confidence: 99%

Mismatch Correction Modulates Mutation Frequency and Pilus Phase and Antigenic Variation in Neisseria gonorrhoeae

et al. 2010

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The mismatch correction (MMC) system repairs DNA mismatches and single nucleotide insertions or deletions postreplication. To test the functions of MMC in the obligate human pathogen Neisseria gonorrhoeae, homologues of the core MMC genes mutS and mutL were inactivated in strain FA1090. No mutH homologue was found in the FA1090 genome, suggesting that gonococcal MMC is not methyl directed. MMC mutants were compared to a mutant in uvrD, the helicase that functions with MMC in Escherichia coli. Inactivation of MMC or uvrD increased spontaneous resistance to rifampin and nalidixic acid, and MMC/uvrD double mutants exhibited higher mutation frequencies than any single mutant. Loss of MMC marginally enhanced the transformation efficiency of DNA carrying a single nucleotide mismatch but not that of DNA with a 1-kb insertion. Unlike the exquisite UV sensitivity of the uvrD mutant, inactivating MMC did not affect survival after UV irradiation. MMC and uvrD mutants exhibited increased PilC-dependent pilus phase variation. mutSdeficient gonococci underwent an increased frequency of pilin antigenic variation, whereas uvrD had no effect. Recombination tracts in the mutS pilin variants were longer than in parental gonococci but utilized the same donor pilS loci. These results show that gonococcal MMC repairs mismatches and small insertion/deletions in DNA and also affects the recombination events underlying pilin antigenic variation. The differential effects of MMC and uvrD in gonococci unexpectedly reveal that MMC can function independently of uvrD in this human-specific pathogen.

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“…The DNA substrates used in the assay are listed in Table 2 . Purified DNA glycosylase Fpg [ 56 ] and bovine serum albumin (BSA) were used as positive and negative controls, respectively. Each experiment was repeated at least three times.…”

Section: Methodsmentioning

confidence: 99%

The Inner Membrane Protein PilG Interacts with DNA and the Secretin PilQ in Transformation

et al. 2015

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Expression of type IV pili (Tfp), filamentous appendages emanating from the bacterial surface, is indispensable for efficient neisserial transformation. Tfp pass through the secretin pore consisting of the membrane protein PilQ. PilG is a polytopic membrane protein, conserved in Gram-positive and Gram-negative bacteria, that is required for the biogenesis of neisserial Tfp. PilG null mutants are devoid of pili and non-competent for transformation. Here, recombinant full-length, truncated and mutated variants of meningococcal PilG were overexpressed, purified and characterized. We report that meningococcal PilG directly binds DNA in vitro, detected by both an electromobility shift analysis and a solid phase overlay assay. PilG DNA binding activity was independent of the presence of the consensus DNA uptake sequence. PilG-mediated DNA binding affinity was mapped to the N-terminus and was inactivated by mutation of residues 43 to 45. Notably, reduced meningococcal transformation of DNA in vivo was observed when PilG residues 43 to 45 were substituted by alanine in situ, defining a biologically significant DNA binding domain. N-terminal PilG also interacted with the N-terminal region of PilQ, which previously was shown to bind DNA. Collectively, these data suggest that PilG and PilQ in concert bind DNA during Tfp-mediated transformation.

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Characterization of the meningococcal DNA glycosylase Fpg involved in base excision repair

Cited by 10 publications

References 45 publications

A network of enzymes involved in repair of oxidative DNA damage in Neisseria meningitidis

A network of enzymes involved in repair of oxidative DNA damage in Neisseria meningitidis

Mismatch Correction Modulates Mutation Frequency and Pilus Phase and Antigenic Variation in Neisseria gonorrhoeae

The Inner Membrane Protein PilG Interacts with DNA and the Secretin PilQ in Transformation

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