title: Comparative analysis of Staphylococcus epidermidis strains utilizing quantitative and cell surface shaving proteomics.
abstract: Staphylococcus epidermidis is an opportunistic pathogen that is an emerging risk factor in hospitals worldwide and is often difficult to eradicate as virulent strains produce a protective biofilm matrix. We utilized cell shaving proteomics to profile surface-exposed proteins from two fully genome sequenced S. epidermidis strains: the avirulent, non-biofilm forming ATCC12228 and the virulent, strongly adherent biofilm forming ATCC35984 (RP62A). A false positive control strategy was employed to calculate the probabilities of proteins being truly surface-exposed. A total of 78 surface-exposed proteins were identified, of which only 19 proteins were common to ATCC12228 and RP62A, and which thus represents the core surfaceome. S. epidermidis RP62A displayed additional proteins involved in biofilm formation (cell wall-associated Bhp and intercellular adhesion protein IcaB), surface antigenicity, peptidoglycan biosynthesis and antibiotic resistance. We concurrently profiled whole cell proteomes of the two strains using iTRAQ quantitation and LC-MS/MS. A total of 1610 proteins were confidently identified (representing 64% of the theoretical S. epidermidis proteome). One hundred and ninety one proteins were differentially abundant between strains. Proteins associated with RP62A were clustered into functions including Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-mediated defense, sulfate assimilation, antibiotic resistance and biofilm formation. Validation of the sulfate assimilation and cysteine/methionine biosynthesis pathways showed RP62A contained elevated levels (~25% increase) of methionine that are likely linked to biofilm formation.
  
  
  CONCLUSIONS
  Cell shaving and quantitative proteomics identified proteins associated with a biofilm-forming, virulent strain of S. epidermidis (RP62A). These proteins show RP62A maintains an active CRISPR-mediated defense, as well as heightened antibiotic resistance in comparison to a non-virulent, non-biofilm forming strain. Increased abundances of sulfate assimilation proteins lead to elevated intracellular methionine. Proteins and their exposed peptides identified on the surface of S. epidermidis RP62A may be useful vaccine antigens in clinical settings if administered in at-risk patients prior to surgical implantations.
authors.affiliation: School of Molecular Bioscience, The University of Sydney 2006, Australia.
authors.forename: Nestor
authors.initials: N
authors.lastname: Solis
authors.affiliation: School of Molecular Bioscience, The University of Sydney 2006, Australia; Discipline of Pathology, School of Medical Sciences, The University of Sydney 2006, Australia.
authors.forename: Joel A
authors.initials: JA
authors.lastname: Cain
authors.affiliation: School of Molecular Bioscience, The University of Sydney 2006, Australia; Discipline of Pathology, School of Medical Sciences, The University of Sydney 2006, Australia; Charles Perkins Centre, The University of Sydney 2006, Australia. Electronic address: stuart.cordwell@sydney.edu.au.
authors.forename: Stuart J
authors.initials: SJ
authors.lastname: Cordwell
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry; D018441:Biofilms; D064113:CRISPR-Cas Systems; D002462:Cell Membrane / Q000378:metabolism*; D024881:Drug Resistance, Bacterial; D005001:Ethionine / Q000737:chemistry; D013058:Mass Spectrometry; D008715:Methionine / Q000737:chemistry; D010641:Phenotype; D011134:Polysaccharides / Q000737:chemistry; D020543:Proteome / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D013045:Species Specificity; D013212:Staphylococcus epidermidis / Q000378:metabolism*; D013431:Sulfates / Q000737:chemistry; D053719:Tandem Mass Spectrometry; D014357:Trypsin / Q000737:chemistry; D014774:Virulence
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D011134:Polysaccharides; D020543:Proteome; D013431:Sulfates; D008715:Methionine; D014357:Trypsin; D005001:Ethionine
keywords: Cell shaving; Mass spectrometry; Methionine biosynthesis; S. epidermidis; Sulfate assimilation; Surfaceome