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Mol Cell Proteomics. 2010 May;9(5):912-27. doi: 10.1074/mcp.M000032-MCP201. Epub 2010 Mar 20.

A statistics-based platform for quantitative N-terminome analysis and identification of protease cleavage products.

Author information

1
Department of Biochemistry and Molecular Biology, Centre for Blood Research, University of British Columbia, 4.401 Life Sciences Institute, 2350 Health Sciences Mall, Vancouver, British Columbia V6T 1Z3, Canada.

Abstract

Terminal amine isotopic labeling of substrates (TAILS), our recently introduced platform for quantitative N-terminome analysis, enables wide dynamic range identification of original mature protein N-termini and protease cleavage products. Modifying TAILS by use of isobaric tag for relative and absolute quantification (iTRAQ)-like labels for quantification together with a robust statistical classifier derived from experimental protease cleavage data, we report reliable and statistically valid identification of proteolytic events in complex biological systems in MS2 mode. The statistical classifier is supported by a novel parameter evaluating ion intensity-dependent quantification confidences of single peptide quantifications, the quantification confidence factor (QCF). Furthermore, the isoform assignment score (IAS) is introduced, a new scoring system for the evaluation of single peptide-to-protein assignments based on high confidence protein identifications in the same sample prior to negative selection enrichment of N-terminal peptides. By these approaches, we identified and validated, in addition to known substrates, low abundance novel bioactive MMP-2 targets including the plasminogen receptor S100A10 (p11) and the proinflammatory cytokine proEMAP/p43 that were previously undescribed.

PMID:
20305283
PMCID:
PMC2871423
DOI:
10.1074/mcp.M000032-MCP201
[Indexed for MEDLINE]
Free PMC Article

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