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Molecular & Cellular Proteomics 6:1188-1197, 2007.
© 2007 by The American Society for Biochemistry and Molecular Biology, Inc.

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Research

Neuropeptidomics Strategies for Specific and Sensitive Identification of Endogenous Peptides^*,S

Maria Fälth^,, Karl Sköld^,, Marcus Svensson^,, Anna Nilsson^,, David Fenyö^¶ and Per E. Andren^,,||

From the Laboratory for Biological and Medical Mass Spectrometry, Biomedical Centre, Box 583, Uppsala University, SE-75123 Uppsala, Sweden, Department of Pharmaceutical Biosciences, Biomedical Centre, Uppsala University, SE-75124 Uppsala, Sweden, and ^¶ The Rockefeller University, New York, New York 10021

A new approach using targeted sequence collections has been developed for identifying endogenous peptides. This approach enables a fast, specific, and sensitive identification of endogenous peptides. Three different sequence collections were constituted in this study to mimic the peptidomic samples: SwePep precursors, SwePep peptides, and SwePep predicted. The searches for neuropeptides performed against these three sequence collections were compared with searches performed against the entire mouse proteome, which is commonly used to identify neuropeptides. These four sequence collections were searched with both Mascot and X! Tandem. Evaluation of the sequence collections was achieved using a set of manually identified and previously verified peptides. By using the three new sequence collections, which more accurately mimic the sample, 3 times as many peptides were significantly identified, with a false-positive rate below 1%, in comparison with the mouse proteome. The new sequence collections were also used to identify previously uncharacterized peptides from brain tissue; 27 previously uncharacterized peptides and potentially bioactive neuropeptides were identified. These novel peptides are cleaved from the peptide precursors at sites that are characteristic for prohormone convertases, and some of them have post-translational modifications that are characteristic for neuropeptides. The targeted protein sequence collections for different species are publicly available for download from SwePep.

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