title: Functional analysis of paralogous thiol-disulfide oxidoreductases in Streptococcus gordonii.
abstract: Disulfide bonds are important for the stability of many extracellular proteins, including bacterial virulence factors. Formation of these bonds is catalyzed by thiol-disulfide oxidoreductases (TDORs). Little is known about their formation in Gram-positive bacteria, particularly among facultative anaerobic Firmicutes, such as streptococci. To investigate disulfide bond formation in Streptococcus gordonii, we identified five putative TDORs from the sequenced genome. Each of the putative TDOR genes was insertionally inactivated with an erythromycin resistance cassette, and the mutants were analyzed for autolysis, extracellular DNA release, biofilm formation, bacteriocin production, and genetic competence. This analysis revealed a single TDOR, SdbA, which exhibited a pleiotropic mutant phenotype. Using an in silico analysis approach, we identified the major autolysin AtlS as a natural substrate of SdbA and showed that SdbA is critical to the formation of a disulfide bond that is required for autolytic activity. Analysis by BLAST search revealed homologs to SdbA in other Gram-positive species. This study provides the first in vivo evidence of an oxidoreductase, SdbA, that affects multiple phenotypes in a Gram-positive bacterium. SdbA shows low sequence homology to previously identified oxidoreductases, suggesting that it may belong to a different class of enzymes. Our results demonstrate that SdbA is required for disulfide bond formation in S. gordonii and indicate that this enzyme may represent a novel type of oxidoreductase in Gram-positive bacteria.
authors.affiliation: Department of Microbiology and Immunology, Dalhousie University, Halifax, Nova Scotia B3H 1X5, Canada; Canadian Center for Vaccinology, Dalhousie University and the Izaak Walton Killam Health Centre, Halifax, Nova Scotia B3K 6R8, Canada.
authors.forename: Lauren
authors.initials: L
authors.lastname: Davey
authors.affiliation: Department of Microbiology and Immunology, Dalhousie University, Halifax, Nova Scotia B3H 1X5, Canada; Canadian Center for Vaccinology, Dalhousie University and the Izaak Walton Killam Health Centre, Halifax, Nova Scotia B3K 6R8, Canada.
authors.forename: Crystal K W
authors.initials: CKW
authors.lastname: Ng
authors.affiliation: Department of Microbiology and Immunology, Dalhousie University, Halifax, Nova Scotia B3H 1X5, Canada; Canadian Center for Vaccinology, Dalhousie University and the Izaak Walton Killam Health Centre, Halifax, Nova Scotia B3K 6R8, Canada; Department of Pediatrics, Faculty of Medicine, Dalhousie University and the Izaak Walton Killam Health Centre, Halifax, Nova Scotia B3K 6R8, Canada.
authors.forename: Scott A
authors.initials: SA
authors.lastname: Halperin
authors.affiliation: Department of Microbiology and Immunology, Dalhousie University, Halifax, Nova Scotia B3H 1X5, Canada; Canadian Center for Vaccinology, Dalhousie University and the Izaak Walton Killam Health Centre, Halifax, Nova Scotia B3K 6R8, Canada; Department of Pediatrics, Faculty of Medicine, Dalhousie University and the Izaak Walton Killam Health Centre, Halifax, Nova Scotia B3K 6R8, Canada; Department of Applied Oral Sciences, Faculty of Dentistry, Dalhousie University, Halifax, Nova Scotia B3H 4R2, Canada. Electronic address: song.lee@dal.ca.
authors.forename: Song F
authors.initials: SF
authors.lastname: Lee
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004220:Disulfides / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000235:genetics / Q000378:metabolism; D011490:Protein Disulfide Reductase (Glutathione) / Q000235:genetics / Q000378:metabolism*; D054773:Streptococcus gordonii / Q000201:enzymology* / Q000235:genetics; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004220:Disulfides; D008565:Membrane Proteins; C105449:SdbA protein, bacteria; D037521:Virulence Factors; D011490:Protein Disulfide Reductase (Glutathione); D009238:N-Acetylmuramoyl-L-alanine Amidase
keywords: Autolysin; Disulfide Bond; Gram-positive; Microbiology; Oxidation-Reduction; Protein Folding; Streptococcus; Streptococcus gordonii; Thiol; Thiol-disulfide Oxidoreductase