Comparison of proteins expressed by Pseudomonas aeruginosa strains representing initial and chronic isolates from a cystic fibrosis patient: an analysis by 2-D gel electrophoresis and capillary column liquid chromatography-tandem mass spectrometry

Isolates of Pseudomonas aeruginosa from chronic lung infections in cystic f ibrosis (CF) patients have phenotypes distinct from those initially infecti ng CF patients, as well as from other clinical or environmental isolates, T o gain a better understanding of the differences in these isolates, protein expression was followed using two-dimensional (2-D) gel electrophoresis an d protein identification by peptide sequencing using micro-capillary column liquid chromatography-tandem mass spectrometry (mu LC/MS/MS). The isolates selected for this analysis were from the sputum of a CF patient: strain 38 3 had a nonmucoid phenotype typical of isolates from the environment, and s train 2192, obtained from the same patient, had a mucoid phenotype typical of isolates from chronic CF lung infections, Strains 383 and 2192 were conf irmed to be genetically identical by restriction endonuclease analysis, ran dom amplified polymorphic DNA-PCR, and pulsed-field gel electrophoresis, Co nditions of protein extraction were optimized for consistent high-resolutio n separation of several hundred proteins from these clinical isolates as de tected by Coomassie staining of 2-D gels, Fourteen proteins were selected f or analysis; this group included those whose expression was common between both strains as well as unique for each strain, The proteins were identifie d by mu LC/MS/MS of the peptides produced by an in-gel tryptic digestion an d compared to translated data from the Pseudomonas Genome Project; optimiza tion of this technique has allowed for the comparison of proteins expressed by strains 383 and 2192.