Abstract
Currently, proteomic tools are able to establish a complete list of the most abundant proteins present in a sample, providing the opportunity to study at high resolution the physiology of any bacteria for which the genome sequence is available. For a comprehensive list, proteins should be first resolved into fractions that are then proteolyzed by trypsin. The resulting peptide mixtures are analyzed by a high-throughput tandem mass spectrometer that records thousands of MS/MS spectra for each fraction. These spectra are then assigned to peptides, which are used as evidence of the existence of proteins. In addition to generating a list of protein identifications, this shortcut to proteomics uses the number of spectra recorded for each protein to quantify the observations. Here, we describe one of the most simple sample preparation methods for high-throughput proteomics of bacteria, as well as the subsequent data processing to extract quantitative information based on the spectral count approach.
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Acknowledgements
This work was supported by the Commissariat à l’Energie Atomique et aux Energies Alternatives, and the Agence Nationale de la Recherche (ANR-12-BSV6-0012-01). E.M.H. was supported by a Fulbright grant. We thank our close colleagues, Anne-Hélène Davin, Guylaine Miotello, Philippe Guérin, Céline Bland, Emie Durighello, Béatrice Alonso, Véronique Malard, and Alain Dedieu, for stimulating discussions.
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Hartmann, E.M., Allain, F., Gaillard, JC., Pible, O., Armengaud, J. (2014). Taking the Shortcut for High-Throughput Shotgun Proteomic Analysis of Bacteria. In: Vergunst, A., O'Callaghan, D. (eds) Host-Bacteria Interactions. Methods in Molecular Biology, vol 1197. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1261-2_16
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DOI: https://doi.org/10.1007/978-1-4939-1261-2_16
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