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M.D. Willcox, M. Bandara, P. Sankaridurg, H. Zhu, S. Cordwell; Salicylic Acid Alters the Membrane Proteome of Pseudomonas aeruginosa . Invest. Ophthalmol. Vis. Sci. 2006;47(13):2653.
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NSAIDS have been shown to affect the adhesion of P. aeruginosa to epithelial cells and contact lenses. The aim of this study was to investigate whether growth of P. aeruginosa 6294 in the presence of sub–inhibitory concentrations of salicylic acid alters its membrane proteome.
P. aeruginosa was grown with or without 30mM salicylic acid. After centrifugation to collect cells, cells were lysed with sonication and membranes were precipitated by incubating in carbonate buffer. Proteins were removed from membranes by incubation in buffer containing zwitter ionic detergent and urea. 2D–gel electrophoresis was used to separate membrane proteins. After electrophoresis, proteins were stained with Sypro Ruby. Gels of membrane proteins obtained with or without salicylic acid were compared using PDQuest software. Gel spots showing reproducible n–fold changes in abundance form two of each set of three repeat gels were then subjected to identification. Identification was by trypsin digest of individual spots followed by MALDI–MS and comparison to proteins in the SwissProt database.
A total of 35 spots were differentially expressed. Most proteins were down–regulated in the presence of salicylic acid (77% of the differentially expressed spots). The 35 spots represented 24 distinct proteins. 11 of these proteins were outer membrane proteins/porins (OMP) or probable OMPs. OMPs OprF, mexA and OprG have roles in antibiotic resistance (generally loss indicating more susceptibility) and these were down–regulated. orpD was down–regulated and oprM was up–regulated, which may indicate increased resistance to antibiotics. MexA and OprM function together, and if levels increase resistance to e.g. quinolones occurs. However, mexA and oprM were differentially altered by salicylic acid and preliminary results show that sensitivity to ciprofloxacin was not altered by treatment with salicylic acid. Three pilus/flagella proteins were down–regulated. This pilus/flagella down–regulation in conjunction with down–regulation of OprF is consistent with the inhibition of adhesion of P. aeruginosa by salicylic acid.
Salicylic acid has potent effects on membrane proteins of P. aeruginosa that explain some of its effects on phenotypic characteristics.
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