The Campylobacter concisus BisA protein plays a dual role: oxide-dependent anaerobic respiration and periplasmic methionine sulfoxide repair

Abstract

Campylobacter concisus, an emerging pathogen found throughout the human oral-gastrointestinal tract, is able to grow under microaerobic or anaerobic conditions; in the latter case, N-or S-oxides could be used as terminal electron acceptors (TEAs). Analysis of 23 genome sequences revealed the presence of multiple (at least two, and up to five) genes encoding for putative periplasmic N- or S-oxide reductases (N/SORs), all of which are predicted to harbor a molybdopterin (or tungstopterin)-bis guanine dinucleotide (Mo/W-bisPGD) cofactor. Various N- or S-oxides, including nicotinamide N-oxide (NANO), trimethylamine N-oxide (TMAO), biotin sulfoxide (BSO), dimethyl sulfoxide (DMSO) and methionine sulfoxide (MetO), significantly increased anaerobic growth in two C. concisus intestinal strains (13826 and 51562) but not in the C. concisus oral (type) strain 33237. A collection of mutants was generated to determine each N/SOR substrate specificity. Surprisingly, we found that disruption of a single gene, annotated as "bisA" (present in strains Cc13826 and Cc51562, but not in Cc33237) abolished all N/S-oxide-supported respiration. Furthermore, {Delta}bisA mutants showed increased sensitivity to oxidative stress and displayed cell envelope abnormalities, suggesting BisA plays a role in protein MetO repair. Indeed, purified recombinant CcBisA was able to successfully repair MetO residues on a commercial protein ({beta}-casein), as shown by mass spectrometry. Our results suggest that BisA plays a dual role in C. concisus, by allowing the pathogen to use N/S-oxides as TEAs, and by repairing periplasmic protein-bound MetO residues, therefore essentially being a periplasmic methionine sulfoxide reductase (Msr). This is the first report of a Mo/W-bisPGD-containing Msr enzyme in a pathogen. IMPORTANCEC. concisus is an excellent model organism to study respiration diversity, including anaerobic respiration of physiologically relevant N/S-oxides compounds, such as BSO, DMSO, MetO, NANO, and TMAO. All C. concisus strains harbor at least two, often three, and up to five genes encoding for putative periplasmic Mo/W-bisPGD-containing N/S-oxide reductases. The respective role (substrate specificity) of each enzyme was studied using a mutagenesis approach. One of the N/SOR enzymes, annotated as "BisA", was found to be essential for anaerobic respiration of both N- and S-oxides. Additional phenotypes associated with disruption of the bisA gene included increased sensitivity toward oxidative stress and elongated cell morphology. Furthermore, a biochemical approach confirmed that BisA can repair protein-bound MetO residues. Hence, we propose that BisA plays a role as a periplasmic methionine sulfoxide reductase. This is the first report of a Mo/W-bisPGD-enzyme supporting both N-or S-oxide respiration and protein-bound MetO repair in a pathogen.