All 4 heat shock proteins (HtpG, DnaK, GroEL and PA4352) were elevated in AES-1R compared to both PAO1 and PA14. Five proteins involved in oxidative stress resistance (PA3529, AhpC, PA4880, PA2331 and KatA) were altered in AES-1R, AZD2014 concentration with all except KatA present at increased abundance. Additional smaller functional clusters included the 3 enzymes of the arginine deiminase

pathway (ArcABC) and the ATP synthase alpha and beta subunits. We identified 2 proteins that were expressed from genes only encoded in the AES-1R genome (spots 26 and 43), and a further protein that was not contained within the PAO1 genome (spot 37). Previously hypothetical protein AES_7139 (spots 43 a-e; Figure 1) was the most abundant protein identified on the 2-DE gels of AES-1R and is present in multiple mass and pI variants. Variants exist at two masses, approximately 28 kDa and 16 kDa, with three pI variants at the higher mass (pI 5.2, 5.6, and

6.0), and two pI variants at the lower mass (pI 5.2 and 6.0). We subjected these spots to both MALDI-TOF MS peptide mass mapping and to LC-MS/MS for sequence characterization. We identified 9 peptide sequences that generated 90.8% sequence coverage for the learn more predicted AES-1R gene (Figure 2). All variants generated near identical MALDI-MS spectra, suggesting the unusual migratory pattern on 2-DE gels are due to folding artifacts or poorly reduced Selleckchem Fludarabine disulfide bonds [31–33]. The AES_7139 translated gene sequence is predicted to encode a protein of 16.7 kDa and with a pI of 5.3, suggesting the higher mass variants may be homodimers or artifacts of the gel process. The sequence contains a single cysteine residue through which a disulfide could be formed, however under the reducing conditions used to conduct 2-DE, it is more likely that a gel artifact results in the spot pattern. One of the peptides sequenced by MS/MS selleckchem displayed a non-tryptic N-terminus 8-GTYLFQYAQDKDYVLGVSDEQSGAK-32 (2782.4093

m/z) cleaved between Met-7 and Gly-8 that suggests either N-terminal processing, or that Met-7 is the true N-terminus. We subjected the AES_7139 protein sequence to BLAST search and showed that there is 100% amino acid sequence identity with a hypothetical protein (PA2G_05851) from P. aeruginosa PA2192 (Blastp score 311, query coverage 100%, e-value 2e-83), an isolate from a chronically infected CF patient in Boston. Other matches displayed similarity to ricin B-type lectins, suggesting the protein might be involved in carbohydrate binding. Importantly, however, no other P. aeruginosa genomes within the Swiss-Prot database contained AES_7139 homologs. Figure 2 Predicted protein sequence of a P. aeruginosa AES-1R hypothetical protein ((A); AES_7139; spot 43a-e) characterized by MALDI-MS and LC-MS/MS (B).