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De novo peptide sequencing and quantitative profiling of complex protein mixtures using mass-coded abundance tagging

Nature Biotechnology volume 20, pages 163–170 (2002)Cite this article

Abstract

Proteomic studies require efficient, robust, and practical methods of characterizing proteins present in biological samples. Here we describe an integrated strategy for systematic proteome analysis based on differential guanidination of C-terminal lysine residues on tryptic peptides followed by capillary liquid chromatography–electrospray tandem mass spectrometry. The approach, termed mass-coded abundance tagging (MCAT), facilitates the automated, large-scale, and comprehensive de novo determination of peptide sequence and relative quantitation of proteins in biological samples in a single analysis. MCAT offers marked advantages as compared with previously described methods and is simple, economic, and effective when applied to complex proteomic mixtures. MCAT is used to identify proteins, including polymorphic variants, from complex mixtures and measure variation in protein levels from diverse cell types.

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Figure 1: Overview of MCAT peptide sequencing and determination of relative protein abundance.
Figure 2: MCAT determination of relative protein abundance in complex biological mixtures.
Figure 3: MCAT is reproducible and quantitative.

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Acknowledgements

We thank Hassan Qureshi, Andy Link, Richard Newitt, John Yates III, David Schieltz, Hayes McDonald, and Mike Washburn for providing reagents, technical advice, and/or comments; Matthew Francis for expert technical assistance; Jimmy Eng for generous use of software; and Shiva Amiri, Thanuja Premawaradena, Alia Qureshi-Emili, Duy Mai, and Mike Lindo for programming expertise. This work was supported in part by a grant from the National Science and Engineering Research Council of Canada to A.E.

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  1. Banting and Best Department of Medical Research, Program in Proteomics and Bioinformatics, University of Toronto, Toronto, ON, Canada

    Gerard Cagney & Andrew Emili

  2. Department of Medical Genetics and Microbiology, University of Toronto, Toronto, ON, Canada

    Andrew Emili

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  1. Gerard Cagney
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Correspondence to Andrew Emili.

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G.C. and A.E. have submitted a provisional patent application for the MCAT methodology/process.

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Cagney, G., Emili, A. De novo peptide sequencing and quantitative profiling of complex protein mixtures using mass-coded abundance tagging. Nat Biotechnol 20, 163–170 (2002). https://doi.org/10.1038/nbt0202-163

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