The disulfide oxidoreductase SdbA is active in Streptococcus gordonii using a single C‐terminal cysteine of the CXXC motif |
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| Authors: | Lauren Davey Alejandro Cohen Jason LeBlanc Scott A Halperin Song F Lee |
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| Institution: | 1. Department of Microbiology and Immunology, Dalhousie University, Halifax, NS, Canada;2. Canadian Center for Vaccinology (CCfV), Dalhousie University and the Izaak Walton Killam (IWK) Health Centre, Halifax, NS, Canada;3. Proteomics and Mass Spectrometry Core Facility, Life Sciences Research Institute, Dalhousie University, Halifax, NS, Canada;4. Department of Pathology, Dalhousie University, Halifax, NS, Canada;5. Department of Pathology and Laboratory Medicine, Nova Scotia Health Authority (NSHA), Halifax, NS, Canada;6. Department of Pediatrics, Faculty of Medicine, Dalhousie University and the IWK Health Centre, Halifax, NS, Canada;7. Department of Applied Oral Sciences, Faculty of Dentistry, Dalhousie University, Halifax, NS, Canada |
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| Abstract: | Recently, we identified a novel disulfide oxidoreductase, SdbA, in the oral bacterium Streptococcus gordonii. Disulfide oxidoreductases form disulfide bonds in nascent proteins using a CXXC catalytic motif. Typically, the N‐terminal cysteine interacts with substrates, whereas the C‐terminal cysteine is buried and only reacts with the first cysteine of the motif. In this study, we investigated the SdbA C86P87D88C89 catalytic motif. In vitro, SdbA single cysteine variants at the N or C‐terminal position (SdbAC86P and SdbAC89A) were active but displayed different susceptibility to oxidation, and N‐terminal cysteine was prone to sulfenylation. In S. gordonii, mutants with a single N‐terminal cysteine were inactive and formed unstable disulfide adducts with other proteins. Activity was partially restored by inactivation of pyruvate oxidase, a hydrogen peroxide generator. Presence of the C‐terminal cysteine alone (in the SdbAC86P variant) could complement the ΔsdbA mutant and restore disulfide bond formation in recombinant and natural protein substrates. These results provide evidence that certain disulfide oxidoreductases can catalyze disulfide bond formation using a single cysteine of the CXXC motif, including the buried C‐terminal cysteine. |
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