Comparative proteomics of Staphylococcus aureus and the response of methicillin-resistant and methicillin-sensitive strains to Triton X-100

• Published in: Microbiology (Society for General Microbiology) Vol. 148; no. 9; pp. 2765 - 2781
• Main Authors: Cordwell, Stuart J, Larsen, Martin R, Cole, Rebecca T, Walsh, Bradley J
• Format: Journal Article
• Language: English
• Published: Reading Soc General Microbiol 01.09.2002; Society for General Microbiology
• Subjects: Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacteriology; Biological and medical sciences; DNA, Bacterial - genetics; Electrophoresis, Gel, Two-Dimensional; Fundamental and applied biological sciences. Psychology; Genes, Bacterial; Genes, Regulator; Genetics; Genome, Bacterial; Mass Spectrometry; Methicillin Resistance - physiology; Microbiology; Octoxynol - pharmacology; Proteomics; Regulon; SarA protein; Staphylococcus aureus; Staphylococcus aureus - drug effects; Staphylococcus aureus - metabolism; Trans-Activators; Shock; Detergent; β-Lactams; Proteome; Meticillin; Infection; Penicillin derivatives; Resistance; Treatment; Bacteriosis; Bacteria; Micrococcales; Micrococcaceae; Staphylococcal infection; Staphylococcus aureus
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/00221287-148-9-2765

Australian Proteome Analysis Facility, Level 4, Building F7B, Macquarie University, Sydney, Australia2109 1 Author for correspondence: Stuart J. Cordwell. Tel: +61 2 9850 6204. Fax: +61 2 9850 6200. e-mail: scordwell{at}proteome.org.au Proteomics is a powerful tool for analysing differences in gene expression between bacterial strains with alternate phenotypes. Staphylococcus aureus strains are grouped on the basis of their sensitivity to methicillin. Two-dimensional gel electrophoresis was combined with MS to compare the protein profiles of S. aureus strains COL (methicillin-resistant) and 8325 (methicillin-sensitive). Reference mapping via this approach identified 377 proteins that corresponded to 266 distinct ORFs. Amongst these identified proteins were 14 potential virulence factors. The production of 41 ‘hypothetical’ proteins was confirmed, and eight of these appeared to be unique to S. aureus . Strain COL displayed 12 protein spots, which included alkaline-shock protein 23 (Asp23) and cold-shock proteins CspABC, which either were not present in strain 8325 or were present at a significantly lower intensity in this strain. Comparative maps were used to characterize the S. aureus response to treatment with Triton X-100 (TX-100), a detergent that has been shown to reduce methicillin resistance independently of an interaction with the mecA -encoded penicillin-binding protein 2a. In response to growth of the bacteria in the presence of TX-100, 44 protein spots showed altered levels of abundance, and 11 of these spots were found only in COL. The products of genes regulated by B (the alternative sigma factor), including Asp23 and three proteins of unknown function, and SarA (a regulator of virulence genes) were shown to be present at significantly altered levels. SarA production was induced in TX-100-treated cultures. A protein of the B operon, RsbV, was only detected in COL and its production was down-regulated in COL when the strain was treated with TX-100, whereas RsbW was present at reduced levels in both strains. Upon growth of both strains in the presence of TX-100, no effects on the production of the essential methicillin-resistance factor FemA were detected, whereas phosphoglucosamine mutase (GlmM) production was reduced in COL alone. This study suggests that proteins of the B and sarA regulons, as well as other factors, are involved in methicillin resistance in S. aureus . View larger version (83K): [in this window] [in a new window]   Fig. 1. 2-DGE of cellular proteins from S. aureus 8325. (a) Proteins separated using IPG pH 4–7 and (b) IPG pH 6–11 two-dimensional gels. Numbers refer to identifications shown in supplementary data (http://mic.sgmjournals.org). Boxes show the positions of spots only found in strain COL. Broken boxes show the positions of spots found only in cells following their growth in the presence of TX-100. NI, not identified; D suffix, potential dimer; F suffix, protein spot corresponds to a fragment of a larger translated ORF.   Keywords: microbial proteome, two-dimensional gel electrophoresis, mass spectrometry, SarA, B Abbreviations: 2-DGE, two-dimensional gel electrophoresis; IPG, immobilized pH gradient; LTA, lipoteichoic acid; MALDI/TOF, matrix-assisted laser desorption/ionization time-of-flight; TX-100, Triton X-100 a The identifications for the spots shown in Fig. 1 can be found as supplementary data in Microbiology Online (http://mic.sgmjournals.org).