Skip to main content
SpringerLink
Log in

Proteomic Approach to Investigate Pathogenicity and Metabolism of Methicillin-Resistant Staphylococcus aureus

  • Protocol
  • First Online:
Methicillin-Resistant Staphylococcus Aureus (MRSA) Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1085))

Abstract

Over the last two decades, numerous genomes of pathogenic bacteria have been fully sequenced and annotated, while others are continuously being sequenced. To date, the sequences of more than 8,500 whole bacterial genomes are publicly available for research purposes. These efforts in high-throughput sequencing simultaneously to progresses in methods allowing to study whole transcriptome and proteome of bacteria provide the basis of comprehensive understanding of metabolism, adaptability to environment, regulation, resistance pathways, or pathogenicity mechanisms of bacterial pathogens. Staphylococcus aureus is a Gram-positive human pathogen causing a wide variety of infections ranging from benign skin infection to life-threatening diseases. Furthermore, the spreading of multidrug-resistant isolates requiring the use of last barrier drugs has resulted in a particular attention of the medical and scientific community to this pathogen. We describe here proteomic methods to prepare, identify, and analyze protein fractions, which allow studying Staphylococcus aureus on the organism level. Besides evaluation of the whole bacterial transcriptome, this approach might contribute to the development of rapid diagnostic tests and to the identification of new drug targets to improve public health.

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

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Lowy FD (1998) Staphylococcus aureus infections. N Engl J Med 339:520–532

    Article  PubMed  CAS  Google Scholar 

  2. Bronner S, Monteil H, Prevost G (2004) Regulation of virulence determinants in Staphylococcus aureus: complexity and applications. FEMS Microbiol Rev 28:183–200

    Article  PubMed  CAS  Google Scholar 

  3. Foster TJ, Hook M (1998) Surface protein adhesins of Staphylococcus aureus. Trends Microbiol 6:484–488

    Article  PubMed  CAS  Google Scholar 

  4. Que YA, Haefliger JA, Piroth L et al (2005) Fibrinogen and fibronectin binding cooperate for valve infection and invasion in Staphylococcus aureus experimental endocarditis. J Exp Med 201:1627–1635

    Article  PubMed  CAS  Google Scholar 

  5. Moreillon P, Entenza JM, Francioli P et al (1995) Role of Staphylococcus aureus coagulase and clumping factor in pathogenesis of experimental endocarditis. Infect Immun 63:4738–4743

    PubMed  CAS  Google Scholar 

  6. Switalski LM, Patti JM, Butcher W et al (1993) A collagen receptor on Staphylococcus aureus strains isolated from patients with septic arthritis mediates adhesion to cartilage. Mol Microbiol 7:99–107

    Article  PubMed  CAS  Google Scholar 

  7. Supersac G, Piemont Y, Kubina M et al (1998) Assessment of the role of gamma-toxin in experimental endophthalmitis using a hlg-deficient mutant of Staphylococcus aureus. Microb Pathog 24:241–251

    Article  PubMed  CAS  Google Scholar 

  8. O’Callaghan RJ, Callegan MC, Moreau JM et al (1997) Specific roles of alpha-toxin and beta-toxin during Staphylococcus aureus corneal infection. Infect Immun 65:1571–1578

    PubMed  Google Scholar 

  9. Clement S, Vaudaux P, Francois P et al (2005) Evidence of an intracellular reservoir in the nasal mucosa of patients with recurrent Staphylococcus aureus rhinosinusitis. J Infect Dis 192:1023–1028

    Article  PubMed  Google Scholar 

  10. Proctor RA, van Langevelde P, Kristjansson M et al (1995) Persistent and relapsing infections associated with small-colony variants of Staphylococcus aureus. Clin Infect Dis 20:95–102

    Article  PubMed  CAS  Google Scholar 

  11. Vaudaux P, Francois P, Bisognano C et al (2002) Increased expression of clumping factor and fibronectin-binding proteins by hemB mutants of Staphylococcus aureus expressing small colony variant phenotypes. Infect Immun 70:5428–5437

    Article  PubMed  CAS  Google Scholar 

  12. Sinha B, Francois PP, Nusse O et al (1999) Fibronectin-binding protein acts as Staphylococcus aureus invasin via fibronectin bridging to integrin α5β1. Cell Microbiol 1:101–117

    Article  PubMed  CAS  Google Scholar 

  13. Kuroda M, Ohta T, Uchiyama I et al (2001) Whole genome sequencing of methicillin-resistant Staphylococcus aureus. Lancet 357:1225–1240

    Article  PubMed  CAS  Google Scholar 

  14. Baba T, Takeuchi F, Kuroda M et al (2002) Genome and virulence determinants of high virulence community-acquired MRSA. Lancet 359:1819–1827

    Article  PubMed  CAS  Google Scholar 

  15. Gill SR, Fouts DE, Archer GL et al (2005) Insights on evolution of virulence and resistance from the complete genome analysis of an early methicillin-resistant Staphylococcus aureus strain and a biofilm-producing methicillin-resistant Staphylococcus epidermidis strain. J Bacteriol 187:2426–2438

    Article  PubMed  CAS  Google Scholar 

  16. Holden MT, Feil EJ, Lindsay JA et al (2004) Complete genomes of two clinical Staphylococcus aureus strains: evidence for the rapid evolution of virulence and drug resistance. Proc Natl Acad Sci U S A 101:9786–9791

    Article  PubMed  CAS  Google Scholar 

  17. Fitzgerald JR, Sturdevant DE, Mackie SM, Gill SR, Musser JM (2001) Evolutionary genomics of Staphylococcus aureus: insights into the origin of methicillin-resistant strains and the toxic shock syndrome epidemic. Proc Natl Acad Sci U S A 98:8821–8826

    Article  PubMed  CAS  Google Scholar 

  18. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Article  PubMed  CAS  Google Scholar 

  19. Russell RB, Eggleston DS (2000) New roles for structure in biology and drug discovery. Nat Struct Biol 7:928–930

    Article  PubMed  CAS  Google Scholar 

  20. Deshusses JM, Burgess JA, Scherl A et al (2003) Exploitation of specific properties of trifluoroethanol for extraction and separation of membrane proteins. Proteomics 3:1418–1424

    Article  PubMed  CAS  Google Scholar 

  21. Scherl A, Francois P, Converset V et al (2004) Nonredundant mass spectrometry: a strategy to integrate mass spectrometry acquisition and analysis. Proteomics 4:917–927

    Article  PubMed  CAS  Google Scholar 

  22. Angel TE, Aryal UK, Hengel SM et al (2012) Mass spectrometry-based proteomics: existing capabilities and future directions. Chem Soc Rev 41:3912–3928

    Article  PubMed  CAS  Google Scholar 

  23. Zubarev RA, Horn DM, Fridriksson EK et al (2000) Electron capture dissociation for structural characterization of multiply charged protein cations. Anal Chem 72:563–573

    Article  PubMed  CAS  Google Scholar 

  24. Syka JE, Coon JJ, Schroeder MJ et al (2004) Peptide and protein sequence analysis by electron transfer dissociation mass spectrometry. Proc Natl Acad Sci U S A 101:9528–9533

    Article  PubMed  CAS  Google Scholar 

  25. Steen H, Mann M (2004) The ABC’s (and XYZ’s) of peptide sequencing. Nat Rev Mol Cell Biol 5:699–711

    Article  PubMed  CAS  Google Scholar 

  26. Fenn JB, Mann M, Meng CK et al (1989) Electrospray ionization for mass spectrometry of large biomolecules. Science 246:64–71

    Article  PubMed  CAS  Google Scholar 

  27. Zeleny J (1914) The electrical discharge from liquid points, and a hydrostatic method of measuring the electric intensity at their surfaces. Phys Rev 3:69–91

    Article  Google Scholar 

  28. Hu Q, Noll RJ, Li H et al (2005) The Orbitrap: a new mass spectrometer. J Mass Spectrom 40:430–443

    Article  PubMed  CAS  Google Scholar 

  29. Bantscheff M, Lemeer S, Savitski MM (2012) Quantitative mass spectrometry in proteomics: critical review update from 2007 to the present. Anal Bioanal Chem 404:939–965

    Article  PubMed  CAS  Google Scholar 

  30. Ross PL, Marchese J, Huang YN et al (2004) Multiplexed protein quantitation in Saccharomyces cerevisiae using amine-reactive isobaric tagging reagents. Mol Cell Proteomics 3(12):1154–1169

    Article  PubMed  CAS  Google Scholar 

  31. Wilkins MR, Gasteiger E, Sanchez JC et al (1998) Two-dimensional gel electrophoresis for proteome projects: the effects of protein hydrophobicity and copy number. Electrophoresis 19:1501–1505

    Article  PubMed  CAS  Google Scholar 

  32. Rabilloud T, Adessi C, Giraudel A et al (1997) Improvement of the solubilization of proteins in two-dimensional electrophoresis with immobilized pH gradients. Electrophoresis 18:307–316

    Article  PubMed  CAS  Google Scholar 

  33. Adessi C, Miege C, Albrieux C et al (1997) Two-dimensional electrophoresis of membrane proteins: a current challenge for immobilized pH gradients. Electrophoresis 18:127–135

    Article  PubMed  CAS  Google Scholar 

  34. Washburn MP, Wolters D, Yates JRIII (2001) Large-scale analysis of the yeast proteome by multidimensional protein identification technology. Nat Biotechnol 19:242–247

    Article  PubMed  CAS  Google Scholar 

  35. Cargile BJ, Talley DL, Stephenson JL Jr (2004) Immobilized pH gradients as a first dimension in shotgun proteomics and analysis of the accuracy of pI predictability of peptides. Electrophoresis 25:936–945

    Article  PubMed  CAS  Google Scholar 

  36. Singer SJ, Nicolson GL (1972) The fluid mosaic model of the structure of cell membranes. Science 175:720–731

    Article  PubMed  CAS  Google Scholar 

  37. Richardson SH, Hultin HO, Green DE (1963) Structural proteins of membrane systems. Proc Natl Acad Sci U S A 50:821–827

    Article  PubMed  CAS  Google Scholar 

  38. Helenius A, Simons K (1975) Solubilization of membranes by detergents. Biochim Biophys Acta 415:29–79

    Article  PubMed  CAS  Google Scholar 

  39. Brugiere S, Kowalski S, Ferro M et al (2004) The hydrophobic proteome of mitochondrial membranes from Arabidopsis cell suspensions. Phytochemistry 65:1693–1707

    Article  PubMed  CAS  Google Scholar 

  40. Ferro M, Salvi D, Riviere-Rolland H et al (2002) Integral membrane proteins of the chloroplast envelope: identification and subcellular localization of new transporters. Proc Natl Acad Sci U S A 99:11487–11492

    Article  PubMed  CAS  Google Scholar 

  41. Ferro M, Seigneurin-Berny D, Rolland N et al (2000) Organic solvent extraction as a versatile procedure to identify hydrophobic chloroplast membrane proteins. Electrophoresis 21:3517–3526

    Article  PubMed  CAS  Google Scholar 

  42. Blonder J, Conrads TP, Yu LR et al (2004) A detergent- and cyanogen bromide-free method for integral membrane proteomics: application to Halobacterium purple membranes and the human epidermal membrane proteome. Proteomics 4:31–45

    Article  PubMed  CAS  Google Scholar 

  43. Guillet V, Roblin P, Werner S et al (2004) Crystal structure of leucotoxin S component: new insight into the Staphylococcal beta-barrel pore-forming toxins. J Biol Chem 279:41028–41037

    Article  PubMed  CAS  Google Scholar 

  44. Guillet V, Keller D, Prevost G et al (2004) Crystallization and preliminary crystallographic data of a leucotoxin S component from Staphylococcus aureus. Acta Crystallogr D Biol Crystallogr 60:310–313

    Article  PubMed  Google Scholar 

  45. Menestrina G, Dalla SM, Comai M et al (2003) Ion channels and bacterial infection: the case of beta-barrel pore-forming protein toxins of Staphylococcus aureus. FEBS Lett 552:54–60

    Article  PubMed  CAS  Google Scholar 

  46. Wallin E, Von Heijne G (1998) Genome-wide analysis of integral membrane proteins from eubacterial, archaean, and eukaryotic organisms. Protein Sci 7:1029–1038

    Article  PubMed  CAS  Google Scholar 

  47. Scherl A, Francois P, Bento M et al (2005) Correlation of proteomic and transcriptomic profiles of Staphylococcus aureus during the post-exponential phase of growth. J Microbiol Methods 60:247–257

    Article  PubMed  CAS  Google Scholar 

  48. Stoughton RB, Friend SH (2005) How molecular profiling could revolutionize drug discovery. Nat Rev Drug Discov 4:345–350

    Article  PubMed  CAS  Google Scholar 

  49. Berger AB, Vitorino PM, Bogyo M (2004) Activity-based protein profiling: applications to biomarker discovery, in vivo imaging and drug discovery. Am J Pharmacogenomics 4:371–381

    Article  PubMed  CAS  Google Scholar 

  50. Lau AT, He QY, Chiu JF (2003) Proteomic technology and its biomedical applications. Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai) 35:965–975

    Google Scholar 

  51. Vytvytska O, Nagy E, Bluggel M et al (2002) Identification of vaccine candidate antigens of Staphylococcus aureus by serological proteome analysis. Proteomics 2:580–590

    Article  PubMed  CAS  Google Scholar 

  52. Haas G, Karaali G, Ebermayer K et al (2002) Immunoproteomics of Helicobacter pylori infection and relation to gastric disease. Proteomics 2:313–324

    Article  PubMed  CAS  Google Scholar 

  53. Ventura CL, Malachowa N, Hammer CH et al (2010) Identification of a novel Staphylococcus aureus two-component leukotoxin using cell surface proteomics. PLoS One 5:e11634

    Article  PubMed  Google Scholar 

  54. Jones RC, Deck J, Edmondson RD et al (2008) Relative quantitative comparisons of the extracellular protein profiles of Staphylococcus aureus UAMS-1 and its sarA, agr, and sarA agr regulatory mutants using one-dimensional polyacrylamide gel electrophoresis and nanocapillary liquid chromatography coupled with tandem mass spectrometry. J Bacteriol 190:5265–5278

    Article  PubMed  CAS  Google Scholar 

  55. Wolf C, Hochgrafe F, Kusch H et al (2008) Proteomic analysis of antioxidant strategies of Staphylococcus aureus: diverse responses to different oxidants. Proteomics 8:3139–3153

    Article  PubMed  CAS  Google Scholar 

  56. Scherl A, Francois P, Charbonnier Y et al (2006) Exploring glycopeptide resistance in Staphylococcus aureus: a combined proteomics and transcriptomics approach for the identification of resistance related markers. BMC Genomics 7:296

    Article  PubMed  Google Scholar 

  57. Staub I, Sieber SA (2009) Beta-lactam probes as selective chemical-proteomic tools for the identification and functional characterization of resistance associated enzymes in MRSA. J Am Chem Soc 131:6271–6276

    Article  PubMed  CAS  Google Scholar 

  58. Hecker M, Antelmann H, Buttner K et al (2008) Gel-based proteomics of Gram-positive bacteria: a powerful tool to address physiological questions. Proteomics 8:4958–4975

    Article  PubMed  CAS  Google Scholar 

  59. Dupuis A, Hennekinne JA, Garin J et al (2008) Protein standard absolute quantification (PSAQ) for improved investigation of staphylococcal food poisoning outbreaks. Proteomics 8:4633–4636

    Article  PubMed  CAS  Google Scholar 

  60. Enany S, Yoshida Y, Magdeldin S et al (2012) Extensive proteomic profiling of the secretome of European community acquired methicillin resistant Staphylococcus aureus clone. Peptides 37:128–137

    Article  PubMed  CAS  Google Scholar 

  61. Monteiro R, Vitorino R, Domingues P et al (2012) Proteome of a methicillin-resistant Staphylococcus aureus clinical strain of sequence type ST398. J Proteomics 75:2892–2915

    Article  PubMed  CAS  Google Scholar 

  62. Le Marechal C, Jardin J, Jan G et al (2011) Staphylococcus aureus seroproteomes discriminate ruminant isolates causing mild or severe mastitis. Vet Res 42:35

    Article  PubMed  Google Scholar 

  63. Cherkasov A, Hsing M, Zoraghi R et al (2011) Mapping the protein interaction network in methicillin-resistant Staphylococcus aureus. J Proteome Res 10:1139–1150

    Article  PubMed  CAS  Google Scholar 

  64. Shoshan SH, Admon A (2005) Proteomics in cancer vaccine development. Expert Rev Proteomics 2:229–241

    Article  PubMed  CAS  Google Scholar 

  65. Le Naour F (2001) Contribution of proteomics to tumor immunology. Proteomics 1:1295–1302

    Article  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Service of Infectious Diseases, Genomic Research Laboratory, Geneva, Switzerland

    Patrice François

  2. Biomedical Proteomics Research Group, Department of Human Protein Sciences, Swiss Center of Applied Human Toxicology, University Medical Center, Geneva, Switzerland

    Alexander Scherl

  3. Department of Human Protein Sciences, Faculty of Medicine, Geneva University, Department of Genetic and Laboratory Medicine, Geneva University Hospitals, Geneva, Switzerland

    Denis Hochstrasser

  4. Genomic Research Laboratory, Division of Infectious Diseases, Geneva University Hospitals, Geneva, Switzerland

    Jacques Schrenzel

Authors
  1. Patrice François
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Alexander Scherl
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Denis Hochstrasser
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jacques Schrenzel
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Dept. Veterinary & Biomedical Sciences, University of Minnesota College of Veterinary Medicine, St. Paul, Minnesota, USA

    Yinduo Ji

Rights and permissions

Reprints and permissions

Copyright information

© 2014 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

François, P., Scherl, A., Hochstrasser, D., Schrenzel, J. (2014). Proteomic Approach to Investigate Pathogenicity and Metabolism of Methicillin-Resistant Staphylococcus aureus . In: Ji, Y. (eds) Methicillin-Resistant Staphylococcus Aureus (MRSA) Protocols. Methods in Molecular Biology, vol 1085. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-664-1_14

Download citation

  • DOI: https://doi.org/10.1007/978-1-62703-664-1_14

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-663-4

  • Online ISBN: 978-1-62703-664-1

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation