A novel role for the VapB‐2 antitoxin of nontypeable Haemophilus influenzae in genomic stability following oxidative stress

Nontypeable Haemophilus influenzae (NTHi) are small Gram‐negative human‐adapted organisms that can cause a number of infections ranging from chronic bronchitis to otitis media. Toxin‐antitoxin (TA) loci are highly conserved gene pairs that are important for mediating growth rates following stress. The function of the NTHi vapBC‐2 TA allele has not yet been characterized; however, gene prediction software placed the VapB‐2 antitoxin in the same group as the Fpg DNA glycosylase, MutM. For an in vivo analysis of VapB‐2 antitoxin DNA glycosylase activity, the vapB‐2 gene was tested for the ability to complement two Escherichia coli mutant strains, JW3610‐2 Δ mutM and JW2926‐1 Δ mutY , following H 2 O 2 ‐induced oxidative stress. The NTHi vapB‐2 gene complemented both the Δ mutM and Δ mutY strains, and there was no significant difference in survival between the wild type strain and the two mutants with vapB‐2 . The VapB‐2 protein was then purified for in vitro assays to characterize its putative DNA glycosylase function. VapB‐2 will be analyzed for its ability to remove 8‐oxodeoxyguanine (8‐oxodG) from a pair with cytosine or to cleave adenine from a pair with 8‐oxodG in 23 base pair substrates in the presence or absence of extracts from Δ mutM or Δ mutY strains. Preliminary data supports a novel role for NTHi VapB‐2 in maintenance of genomic stability following oxidative stress. Supported by NIH R01‐DC010187.