Proteomics has recently demonstrated utility in understanding cellular processes on the molecular level as a component of systems biology approaches and for identifying potential biomarkers of various disease states. The large amount of data generated by utilizing high efficiency (e.g., chromatographic) separations coupled to high mass accuracy mass spectrometry for high-throughput proteomics analyses presents challenges related to data processing, analysis, and display. This review focuses on recent advances in nanoLC-FTICR-MS-based proteomics approaches and the accompanying data processing tools that have been developed to display and interpret the large volumes of data being produced.

The pathogen Salmonella enterica is known to cause both food poisoning and typhoid fever. Because of the emergence of antibiotic-resistant isolates and the threat of bioterrorism (e.g. contamination of the food supply), there is a growing need to study this bacterium. In this investigation, comparative peptidomics was used to study S. enterica serovar Typhimurium cultured in either a rich medium or in an acidic, low magnesium, and minimal nutrient medium designed to roughly mimic the macrophage phagosomal environment (within which Salmonella are known to survive). Native peptides from cleared cell lysates were enriched by using isopropanol extraction and analyzed by using both LC-MS/MS and LC-FTICR-MS. We identified and quantified 5,163 peptides originating from 682 proteins, and the data clearly indicated that compared with Salmonella cultured in the rich medium, cells cultured in the phagosome-mimicking medium had dramatically higher abundances of a wide variety of protein degradation products, especially from ribosomal proteins. Salmonella from the same cultures were also analyzed using traditional, bottom-up proteomic methods, and when the peptidomics and proteomics data were analyzed together, two clusters of proteins targeted for proteolysis were tentatively identified. Possible roles of targeted proteolysis by phagocytosed Salmonella are discussed.

Leukotriene A 4 (LTA 4 ) is a chemically unstable triene epoxide product of 5-lipoxygenase metabolism of arachidonic acid. Despite this chemical reactivity and its synthesis at the perinuclear membrane, LTA 4 is enzymatically converted into the cysteinyl leukotrienes and leukotriene B 4 . Furthermore, LTA 4 participates in transcellular biosynthesis and is thus transferred between cells as an intact molecule. A cytosolic fatty acid-binding protein present in the rat basophilic leukemia cells was identified using mass spectrometry. This protein was determined to be the stabilizing factor present in the cell cytosol responsible for increasing the effective chemical half-life of LTA 4 . Rat epithelial fatty acid-binding protein (E-FABP) was isolated using partial protein purification and immunoprecipitation. In-gel digestion with trypsin followed by peptide fingerprint analysis using matrix-assisted laser desorption ionization mass spectrometry and sequencing the major tryptic peptide obtained from liquid chromatography/mass spectrometry/mass spectrometry analysis identified E-FABP in the active fraction. Semi-quantitative Western blot analysis indicated that E-FABP in the cytosolic fraction of RBL-1 cells was present at ϳ1-3 pmol/10 6 cells. E-FABP (9 M) was tested for its ability to stabilize LTA 4 , and at 37°C E-FABP was able to increase the half-life of LTA 4 from the previously reported half-life less than 3 s to a halflife of ϳ7 min. These results present a novel function for the well studied fatty acid-binding protein as a participant in leukotriene biosynthesis that permits LTA 4 to be available for further enzymatic processing in various cellular regions.

DBS techniques for the bioanalysis of drugs and metabolites from whole blood have been demonstrated to be a useful tool in drug development. The term dried matrix spot (DMS) has been used to indicate that the DBS technique has been applied to nonblood matrices. DMS methods often employ a color-indicating process that enhances the ability to analyze these mostly transparent fluids when spotted onto collection paper. The color-indicating dye allows the analyst to visually confirm the location of the dried sample spot. Other benefits of using a color-indicating dye include improved method accuracy and precision, because the process of adding the dye allows for the concurrent addition of the IS prior to sample addition and extraction. To date, matrices that have been analyzed using DMS include cerebrospinal fluid, synovial fluid, saliva, tears, urine and plasma.