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Digestion and depletion of abundant proteins improves proteomic coverage

Nature Methods volume 10, pages 5456 (2013) | Download Citation

  • An Addendum to this article was published on 27 February 2014

Abstract

Two major challenges in proteomics are the large number of proteins and their broad dynamic range in the cell. We exploited the abundance-dependent Michaelis-Menten kinetics of trypsin digestion to selectively digest and deplete abundant proteins with a method we call DigDeAPr. We validated the depletion mechanism with known yeast protein abundances, and we observed greater than threefold improvement in low-abundance human-protein identification and quantitation metrics. This methodology should be broadly applicable to many organisms, proteases and proteomic pipelines.

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Acknowledgements

This project was supported by the US National Center for Research Resources (5P41RR011823-17), National Institute of General Medical Sciences (8P41GM103533-17), National Institute of Diabetes and Digestive and Kidney Diseases (R01DK074798), National Heart, Lung, and Blood Institute (RFP-NHLBI-HV-10-5) and National Institute of Mental Health (R01MH067880). We thank J.N. Savas, C.M. Delahunty and J.K. Diedrich for comments on the manuscript.

Author information

Affiliations

  1. Department of Chemical Physiology, The Scripps Research Institute, La Jolla, California, USA.

    • Bryan R Fonslow
    • , Benjamin D Stein
    • , Kristofor J Webb
    • , Tao Xu
    • , Jeong Choi
    • , Sung Kyu Park
    •  & John R Yates III

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Contributions

B.R.F. designed experiments, performed experiments, analyzed data and wrote the manuscript. B.D.S. prepared HEK cell lysates and provided conceptual advice. K.J.W. prepared yeast lysates. T.X., J.C. and S.K.P. developed software for data analysis. J.R.Y. wrote the manuscript and provided conceptual guidance.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to John R Yates III.

Supplementary information

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    Supplementary Text and Figures

    Supplementary Figures 1–8 and Supplementary Notes 1–6

Excel files

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    Supplementary Data

    Protein and peptide raw data and comparisons from yeast and HEK-cell control and DigDeAPr analyses.

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DOI

https://doi.org/10.1038/nmeth.2250

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