U.S. Department of Energy

Pacific Northwest National Laboratory

Performing comparative peptidomics analyses of Salmonella from different growth conditions.

TitlePerforming comparative peptidomics analyses of Salmonella from different growth conditions.
Publication TypeJournal Article
Year of Publication2010
AuthorsAdkins JN, Mottaz H, Metz TO, Ansong C, Manes NP, Smith RD, Heffron F
JournalMethods Mol Biol
KeywordsAmino Acid Sequence, Animals, Bacterial Proteins, Chromatography, Liquid, Cluster Analysis, Mice, Molecular Sequence Data, Peptides, Proteomics, Salmonella typhimurium, Tandem Mass Spectrometry
Abstract

Host-pathogen interactions are complex competitions during which both the host and the pathogen adapt rapidly to each other in order for one or the other to survive. Salmonella enterica serovar Typhimurium is a pathogen with a broad host range that causes a typhoid fever-like disease in mice and severe food poisoning in humans. The murine typhoid fever is a systemic infection in which S. typhimurium evades part of the immune system by replicating inside macrophages and other cells. The transition from a foodborne contaminant to an intracellular pathogen must occur rapidly in multiple, ordered steps in order for S. typhimurium to thrive within its host environment. Using S. typhimurium isolated from rich culture conditions and from conditions that mimic the hostile intracellular environment of the host cell, a native low molecular weight protein fraction, or peptidome, was enriched from cell lysates by precipitation of intact proteins with organic solvents. The enriched peptidome was analyzed by both LC-MS/MS and LC-MS-based methods, although several other methods are possible. Pre-fractionation of peptides allowed identification of small proteins and protein degradation products that would normally be overlooked. Comparison of peptides present in lysates prepared from Salmonella grown under different conditions provided a unique insight into cellular degradation processes as well as identification of novel peptides encoded in the genome but not annotated. The overall approach is detailed here as applied to Salmonella and is adaptable to a broad range of biological systems.

DOI10.1007/978-1-60761-535-4_2
Alternate JournalMethods Mol. Biol.
PubMed ID20013197
Grant ListR01 AI022933-24 / AI / NIAID NIH HHS / United States
R01 AI022933-25 / AI / NIAID NIH HHS / United States
RR18522 / RR / NCRR NIH HHS / United States
Y1-AI-4894-01 / AI / NIAID NIH HHS / United States
Y1-AI-8401-01 / AI / NIAID NIH HHS / United States
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