Characterisation of the outer membrane proteome of Salmonella Spp

Despite successful sequencing and subsequent comparison of Salmonella Typhimurium (S. Typhimurium) and Salmonella Typhi (S. Typhi) genomes, mechanisms driving S. Typhi host specificity and pathogenicity remain unclear. The characterisation of the surface proteomes of the two serotypes may reveal proteins that play a role in host pathogen interaction, thereby permitting S. Typhi to establish persistent infections. Surface proteins represent a small fraction of the total cell and are difficult to extract preferentially. This study evaluated a range of methods for the extraction of outer membrane proteins (OMPs) in S. Typhimurium, including previously published protocols based on differential solubilisation and a novel approach based on the LPI™ FlowCell. Liquid chromatography tandem mass spectrometry (LC-MS/MS) identified these proteins, and their association with the outer membrane confirmed against a database containing in silica predictions of the sub-cellular location of all ORFs in both genomes. This comprehensive approach identified 73 OMPs, representing 63% of all predicted S. Typhimurium aMPs. When applied to S. Typhi, several OMPs undetected in S. Typhimurium extracts were identified including proteins involved in capsule synthesis. Two-dimensional polyacrylamide gel electrophoresis (2D PAGE) detected OMPs potentially upregulated in S. Typhi, resulting in the identification of 20 upregulated spots containing 15 OMPs, five previously undetected. OMPs with various functions were identified, including porins, receptor proteins, transporters and enzymes. Several OMPs with potential roles in virulence, such as the two regulators SlyB and YfgL modulati ng the expression of pathogenicity islands-encoded proteins, were detected. Additionally, the TolC protein previously implicated in antibiotic resistance and invasion as part of the AcrAB-ToIC efflux system was present in both serotypes. Other examples include the FepA and IroN, involved in iron uptake, potentially upregulated in S. Typhi. The expression of many hypothetical aMPs with poorly defined functions was also confirmed, providing useful targets for future studies.EThOS - Electronic Theses Online ServiceGBUnited Kingdo