Skip to main content
Log in

Staphylococcus aureus DsbA is a membrane-bound lipoprotein with thiol-disulfide oxidoreductase activity

  • Original Paper
  • Published:
Archives of Microbiology Aims and scope Submit manuscript

Abstract

DsbA proteins, the primary catalysts of protein disulfide bond formation, are known to affect virulence and penicillin resistance in Gram-negative bacteria. We identified a putative DsbA homologue in the Gram-positive pathogen Staphylococcus aureus that was able to restore the motility phenotype of an Escherichia coli dsbA mutant and thus demonstrated a functional thiol oxidoreductase activity. The staphylococcal DsbA (SaDsbA) had a strong oxidative redox potential of −131 mV. The persistence of the protein throughout the growth cycle despite its predominant transcription during exponential growth phase suggested a rather long half-life for the SaDsbA. SaDsbA was found to be a membrane localised lipoprotein, supporting a role in disulfide bond formation. But so far, neither in vitro nor in vivo phenotype could be identified in a staphylococcal dsbA mutant, leaving its physiological role unknown. The inability of SaDsbA to interact with the E. coli DsbB and the lack of an apparent staphylococcal DsbB homologue suggest an alternative re-oxidation pathway for the SaDsbA.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

References

  • Akman L, Yamashita A, Watanabe H, Oshima K, Shiba T, Hattori M, Aksoy S, Yamashita A, Watanabe H, Oshima K, Shiba T, Hattori M, Aksoy S (2002) Genome sequence of the endocellular obligate symbiont of tsetse flies, Wigglesworthia glossinidia. Nat Genet 32:402–407

    Article  PubMed  CAS  Google Scholar 

  • Altschul S, Gish W, Miller W, Myers E, Lipman D (1990) Basic local alignment search tool. J Mol Biol 215:403–410

    Article  PubMed  CAS  Google Scholar 

  • Anfinsen CB (1973) Principles that govern the folding of protein chains. Science 181:223–230

    Article  PubMed  CAS  Google Scholar 

  • Bader M, Muse W, Zander T, Bardwell J (1998) Reconstitution of a protein disulfide catalytic system. J Biol Chem 273:10302–10307

    Article  PubMed  CAS  Google Scholar 

  • Bader M, Winther JR, Bardwell JC (1999) Protein oxidation: prime suspect found ’not guilty’. Nat Cell Biol 1:E57–E58

    Article  PubMed  CAS  Google Scholar 

  • Bardwell J, McGovern K, Beckwith J (1991) Identification of a protein required for disulfide bond formation in vivo. Cell 67:581–589

    Article  PubMed  CAS  Google Scholar 

  • Bendtsen JD, Nielsen H, von Heijne G, Brunak S (2004) Improved prediction of signal peptides: SignalP 3.0. J Mol Biol 340:783–795

    Article  PubMed  CAS  Google Scholar 

  • Berger-Bächi B (1983) Increase in transduction efficiency of Tn 551 mediated by the methicillin resistance marker. J Bacteriol 154:533–535

    PubMed  Google Scholar 

  • Berger-Bächi B, Kohler ML (1983) A novel site on the chromosome of Staphylococcus aureus influencing the level of methicillin resistance: genetic mapping. FEMS Microbiol Lett 20:305–309

    Article  Google Scholar 

  • Bischoff M, Entenza JM, Giachino P (2001) Influence of a functional sigB operon on the global regulators sar and agr in Staphylococcus aureus. J Bacteriol 183:5171–5179

    Article  PubMed  CAS  Google Scholar 

  • Bolhuis A, Venema G, Quax W, Bron S, van Dijl J (1999) Functional analysis of paralogous thiol-disulfide oxidoreductases in Bacillus subtilis. J Biol Chem 274:24531–24538

    Article  PubMed  CAS  Google Scholar 

  • Bolivar F, Rodriguez R, Greene P, Betlach MC, Heyneker HL, Boyer HW (1977) Construction and characterisation of new cloning vehicles II A multipurpose cloning system. Gene 2:95–113

    Article  PubMed  CAS  Google Scholar 

  • Bremell T, Lange S, Yacoub A, Ryden C, Tarkowski A (1991) Experimental Staphylococcus aureus arthritis in mice. Infect Immun 59:2615–2623

    PubMed  CAS  Google Scholar 

  • Brückner R (1997) Gene replacement in Staphylococcus carnosus and Staphylococcus xylosus. FEMS Microbiol Lett 151:1–8

    Article  PubMed  Google Scholar 

  • Burall LS, Harro JM, Li X, Lockatell CV, Himpsl SD, Hebel JR, Johnson DE, Mobley HL (2004) Proteus mirabilis genes that contribute to pathogenesis of urinary tract infection: identification of 25 signature-tagged mutants attenuated at least 100-fold. Infect Immun 72:2922–2938

    Article  PubMed  CAS  Google Scholar 

  • Charbonnier J, Belin P, Moutiez M, Stura EA, Quéméneur E (1999) On the role of the cis-proline residue in the active site of DsbA. Protein Sci 8:96–105

    Article  PubMed  CAS  Google Scholar 

  • Chawla RK, Lewis FW, Kutner MH, Bate DM, Roy RG, Rudman D (1984) Plasma cysteine, cystine, and glutathione in cirrhosis. Gastroenterology 87:770–776

    PubMed  CAS  Google Scholar 

  • Cheung AL, Eberhardt KJ, Fischetti VA (1994) A method to isolate RNA from Gram-positive bacteria and mycobacteria. Anal Biochem 222:511–514

    Article  PubMed  CAS  Google Scholar 

  • Dailey F, Berg H (1993) Mutants in disulfide bond formation that disrupt flagellar assembly in Escherichia coli. Proc Natl Acad Sci USA 90:1043–1047

    Article  PubMed  CAS  Google Scholar 

  • Duthie E, Lorenz L (1952) Staphylococcal coagulase: mode of action and antigenicity. J Gen Microbiol 6:95–107

    PubMed  CAS  Google Scholar 

  • Dziewanowska K, Edwards V, Deringer J, Bohach G, Guerra D (1996) Comparison of the β-toxins from Staphylococcus aureus and Staphylococcus intermedius. Arch Biochem Biophys 335:102–108

    Article  PubMed  CAS  Google Scholar 

  • Erlendsson L, Hederstedt L (2002) Mutations in the thiol-disulfide oxidoreductases BdbC and BdbD can suppress cytochrome c deficiency of CcdA-defective Bacillus subtilis cells. J Bacteriol 184:1423–1429

    Article  PubMed  CAS  Google Scholar 

  • Fabianek R, Hennecke H, Thöny-Meyer L (1998) The active-site cysteines of the periplasmic thioredoxin-like CcmG of Escherichia coli are important but not essential for cytochrome c maturation in vivo. J Bacteriol 180:1947–1950

    PubMed  CAS  Google Scholar 

  • Fabianek R, Hennecke H, Thöny-Meyer L (2000) Periplasmic protein thiol:disulphide oxidoreductases of Escherischia coli. FEMS Microbiol Lett 24:303–316

    Article  CAS  Google Scholar 

  • Giachino P, Engelmann S, Bischoff M (2001) σB activity depends on RsbU in Staphylococcus aureus. J Bacteriol 183:1843–1852

    Article  PubMed  CAS  Google Scholar 

  • Grauschopf U, Fritz A, Glockshuber R (2003) Mechanism of the electron transfer catalyst DsbB from Escherichia coli. EMBO J 22:3503–3513

    Article  PubMed  CAS  Google Scholar 

  • Guilhot C, Jander G, Martin N, Beckwith J (1995) Evidence that the pathway of disulfide bond formation in Escherichia coli involves interactions between the cysteines of DsbA and DsbB. Proc Natl Acad Sci USA 92:9895–9899

    Article  PubMed  CAS  Google Scholar 

  • Ha UH, Wang Y, Jin S (2003) DsbA of Pseudomonas aeruginosa is essential for multiple virulence factors. Infect Immun 71:1590–1595

    Article  PubMed  CAS  Google Scholar 

  • van Ham RCHJ, Kamerbeek J, Palacios C, Rausell C, Abascal F, Bastolla U, Fernandez JM, Jimenez L, Postigo M, Silva FJ, Tamames J, Viguera E, Latorre A, Valencia A, Moran F, Moya A (2003) Reductive genome evolution in Buchnera aphidicola. Proc Natl Acad Sci USA 100:581–586

    Article  PubMed  CAS  Google Scholar 

  • Helmann JD (1995) Compilation and analysis of Bacillus subtilis σA -dependent promoter sequences: evidence for extended contact between RNA polymerase and upstream promoter DNA. Nucleic Acid Res 23:2351–2360

    Article  PubMed  CAS  Google Scholar 

  • Hovde CJ, Marr JC, Hoffmann ML, Hackett SP, Chi Y, Crum KK, Stevens DL, Stauffacher CV, Bohach GA (1994) Investigation of the role of the disulphide bond in the activity and structure of staphylococcal enterotoxin C1. Mol Microbiol 13:897–909

    Article  PubMed  CAS  Google Scholar 

  • Huber-Wunderlich M, Glockshuber R (1998) A single dipeptide sequence modulates the redox properties of a whole enzyme family. Fold Des 3:161–171

    Article  PubMed  CAS  Google Scholar 

  • Inukai M, Ghrayeb J, Nakamura K, Inouye M (1984) Apolipoprotein, an intermediate in the processing of the major lipoprotein of the Escherichia coli outer membrane. J Biol Chem 259:757–760

    PubMed  CAS  Google Scholar 

  • Ishihara T, Hideaki T, Hasegawa Y, Tsukagoshi N, Yamagata H, Udaka S (1995) Cloning and characterization of the gene for a protein:thiol-disulfide oxidoreductase in Bacillus brevis. J Bacteriol 177:745–749

    PubMed  CAS  Google Scholar 

  • Jonda S, Huber-Wunderlich M, Glockshuber R, Mossner E (1999) Complementation of DsbA deficiency with secreted thioredoxin variants reveals the crucial role of an efficient dithiol oxidant for catalyzed protein folding in the bacterial periplasm. Embo J 18:3271–3281

    Article  PubMed  CAS  Google Scholar 

  • Jones C, Macnab R, Okino H, Aizawa S (1990) Stoichiometric analysis of the flagellar hook- (basal-body) complex of Salmonella typhimurium. J Mol Biol 212:377–387

    Article  PubMed  CAS  Google Scholar 

  • Jones D, Carlson J, Mody V, Cai J, Lynn M, Sternberg P (2000) Redox state of glutathione in human plasma. Free Radic Biol Med 28:625–635

    Article  PubMed  CAS  Google Scholar 

  • Kadokura H, Katzen F, Beckwith J (2003) Protein disulfide bond formation in prokaryotes. Annu Rev Biochem 72:111–135

    Article  PubMed  CAS  Google Scholar 

  • Kadokura H, Tian H, Zander T, Bardwell J, Beckwith J (2004) Snapshots of DsbA in action: detection of proteins in the process of oxidative folding. Science 303:534–537

    Article  PubMed  CAS  Google Scholar 

  • Kishigami S, Ito K (1996) Roles of cysteine residues of DsbB in its activity to reoxidize DsbA, the protein disulphide bond catalyst of Escherichia coli. Genes Cells 1:201–208

    Article  PubMed  CAS  Google Scholar 

  • Kishigami S, Kanaya E, Kikuchi M, Ito K (1995) DsbA-DsbB interaction through their active site cysteines. J Biol Chem 270:17072–17074

    Article  PubMed  CAS  Google Scholar 

  • Martin JL (1995) Thioredoxin–a fold for all reasons. Structure 3:245–250

    Article  PubMed  CAS  Google Scholar 

  • Meima R, Eschevins C, Fillinger S, Bolhuis A, Hamoen LW, Dorenbos R, Quax WJ, van Dijl JM, Provvedi R, Chen I, Dubnau D, Bron S (2002) The bdbDC operon of Bacillus subtilis encodes thiol-disulfide oxidoreductases required for competence development. J Biol Chem 277:6994–7001

    Article  PubMed  CAS  Google Scholar 

  • Merchante R, Pooley H, Karamata D (1995) A periplasm in Bacillus subtilis. J Bacteriol 177:6176–6183

    PubMed  CAS  Google Scholar 

  • Miki T, Okada N, Danbara H (2004) Two periplasmic disulfide oxidoreductases, DsbA and SrgA, target outer membrane protein SpiA, a component of the Salmonella pathogenicity island 2 type III secretion system. J Biol Chem 279:34631–34642

    Article  PubMed  CAS  Google Scholar 

  • Missiakas D, Georgopoulos C, Raina S (1993) Identification and characterization of the Escherichia coli gene dsbB, whose product is involved in the formation of disulfide bonds in vivo. Proc Natl Acad Sci USA 90:7084–7088

    Article  PubMed  CAS  Google Scholar 

  • Navarre WW, Daefler S, Schneewind O (1996) Cell wall sorting of lipoproteins in Staphylococcus aureus. J Bacteriol 178:441–446

    PubMed  CAS  Google Scholar 

  • Ng T, Kwik J, Maier R (1997) Cloning and expression of the gene for a protein disulfide oxidoreductase from Azotobacter vinelandii: complementation of an Escherichia coli dsbA mutant strain. Gene 188:109–113

    Article  PubMed  CAS  Google Scholar 

  • Nielsen J, Lampen J (1982) Glyceride-cysteine lipoproteins and secretion by Gram-positive bacteria. J Bacteriol 152:315–322

    PubMed  CAS  Google Scholar 

  • Nilsson IM, Lee JC, Bremell T, Ryden C, Tarkowski A (1997) The role of staphylococcal polysaccharide microcapsule expression in septicemia and septic arthritis. Infect Immun 65:4216–4221

    PubMed  CAS  Google Scholar 

  • Peng HL, Novick RP, Kreiswirth B, Kornblum J, Schlievert P (1988) Cloning, characterization, and sequencing of an accessory gene regulator (agr) in Staphylococcus aureus. J Bacteriol 170:4365–4372

    PubMed  CAS  Google Scholar 

  • Pettit RK, Judd RC (1992) The interaction of naturally elaborated blebs from serum-susceptible and serum-resistant strains of Neisseria gonorrhoeae with normal human serum. Mol Microbiol 6:729–734

    Article  PubMed  CAS  Google Scholar 

  • Rao L, Karls RK, Betley MJ (1995) In vitro transcription of pathogenesis-related genes by purified RNA polymerase from Staphylococcus aureus. J Bacteriol 177:2609–2614

    PubMed  CAS  Google Scholar 

  • Rhoads Kraemer G, Iandolo J (1990) High-frequency transformation of Staphylococcus aureus by electroporation. Curr Microbiol 21:373–376

    Article  Google Scholar 

  • Rossi J, Bischoff M, Wada A, Berger-Bächi B (2003) MsrR, a putative cell envelope-associated element involved in Staphylococcus aureus sarA attenuation. Antimicrob Agents Chemother 47:2558–2564

    Article  PubMed  CAS  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (eds) (1989) Molecular cloning, a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory, New York

    Google Scholar 

  • Schierle CF, Berkmen M, Huber D, Kumamoto C, Boyd D, Beckwith J (2003) The DsbA signal sequence directs efficient, cotranslational export of passenger proteins to the Escherichia coli periplasm via the signal recognition particle pathway. J Bacteriol 185:5706–5713

    Article  PubMed  CAS  Google Scholar 

  • Schneewind O, Mihaylova-Petkov D, Model P (1993) Cell wall sorting signals in surface proteins of Gram-positive bacteria. EMBO J 12:4803–4811

    PubMed  CAS  Google Scholar 

  • Sifri CD, Begun J, Ausubel FM, Calderwood SB (2003) Caenorhabditis elegans as a model host for Staphylococcus aureus pathogenesis. Infect Immun 71:2208–2217

    Article  PubMed  CAS  Google Scholar 

  • Stenson TH, Weiss AA (2002) DsbA and DsbC are required for secretion of pertussis toxin by Bordetella pertussis. Infect Immun 70:2297–2303

    Article  PubMed  CAS  Google Scholar 

  • Sutcliffe IC, Harrington DJ (2002) Pattern searches for the identification of putative lipoprotein genes in Gram-positive bacterial genomes. Microbiology 148:2065–2077

    PubMed  CAS  Google Scholar 

  • Thompson J, Higgins D, Gibson T (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680

    Article  PubMed  CAS  Google Scholar 

  • Thöny-Meyer L, Künzler P, Hennecke H (1998) Requirements for maturation of Bradyrhizobium japonicum cytochrome c550 in Escherichia coli. Eur J Biochem 235:754–761

    Article  Google Scholar 

  • Tinsley CR, Voulhoux R, Beretti J-L, Tommassen J, Nassif X (2004) Three homologues, including two membrane-bound proteins, of the disulfide oxidoreductase DsbA in Neisseria meningitidis: effects on bacterial growth and biogenesis of functional type IV pili. J Biol Chem 279:27078–27087

    Article  PubMed  CAS  Google Scholar 

  • Tjalsma H, Bolhuis A, Jongbloed J, Bron S, van Dijl J (2000) Signal peptide-dependent protein transport in Bacillus subtilis: a genome-based survey of the secretome. Microbiol Mol Biol Rev 64:515–547

    Article  PubMed  CAS  Google Scholar 

  • Wada A (2001) An improved method for purifying bacterial genomic DNAs for direct sequencing by capillary automated sequencer. TTO 1:T02049

    Google Scholar 

  • Yu J, Kroll JS (1999) DsbA: a protein-folding catalyst contributing to bacterial virulence. Microbes Infect 1:1221–1228

    Article  PubMed  CAS  Google Scholar 

  • Yu J, Oragui EE, Stephens A, Kroll JS, Venkatesan MM (2001) Inactivation of DsbA alters the behaviour of Shigella flexneri towards murine and human-derived macrophage-like cells. FEMS Microbiol Lett 204:81–88

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We thank J.C.A. Bardwell for providing the strain JCB570 and JCB571, I.M. Jonsson and C. Sifri for performing the animal studies and Sankyo Corp (Tokyo, Japan) for the kind gift of globomycin. This work was supported by a grant from the Olga Mayenfisch Foundation, and the SNF grants 31-105390 and 3100A0-100234.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Brigitte Berger-Bächi.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Dumoulin, A., Grauschopf, U., Bischoff, M. et al. Staphylococcus aureus DsbA is a membrane-bound lipoprotein with thiol-disulfide oxidoreductase activity. Arch Microbiol 184, 117–128 (2005). https://doi.org/10.1007/s00203-005-0024-1

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00203-005-0024-1

Keywords

Navigation