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Global Ubiquitination Analysis by SILAC in Mammalian Cells

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Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC)

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1188))

Abstract

Ubiquitination is a versatile and dynamic posttranslational modification in cells, regulating almost all cellular events. With rapid developments of affinity capture reagents and high-resolution mass spectrometry, it is now feasible to globally analyze the ubiquitinated proteome (ubiquitome) using quantitative strategies, such as stable isotope labeling with amino acids in cell culture (SILAC). Here we describe in detail a SILAC protocol to profile the ubiquitome in mammalian cells including protein labeling, antibody-based enrichment, and analysis by mass spectrometry.

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Acknowledgements

This work was partially supported by National Institutes of Health grant NS081571 and American Cancer Society grant RSG-09-181, and ALSAC (American Lebanese Syrian Associated Charities).

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Authors and Affiliations

  1. Department of Structural Biology, St. Jude Proteomics Facility, St Jude Children’s Research Hospital, Memphis, TN, 38105, USA

    Zhiping Wu, Chan Hyun Na, Haiyan Tan & Junmin Peng

  2. Department of Developmental Neurobiology, St. Jude Proteomics Facility, St Jude Children’s Research Hospital, Memphis, TN, 38105, USA

    Zhiping Wu, Chan Hyun Na, Haiyan Tan & Junmin Peng

Authors
  1. Zhiping Wu
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  2. Chan Hyun Na
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  3. Haiyan Tan
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  4. Junmin Peng
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Corresponding author

Correspondence to Junmin Peng .

Editor information

Editors and Affiliations

  1. Institute of Biology II, University of Freiburg, Freiburg, Germany

    Bettina Warscheid

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© 2014 Springer Science+Business Media New York

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Wu, Z., Na, C.H., Tan, H., Peng, J. (2014). Global Ubiquitination Analysis by SILAC in Mammalian Cells. In: Warscheid, B. (eds) Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC). Methods in Molecular Biology, vol 1188. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1142-4_11

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  • DOI: https://doi.org/10.1007/978-1-4939-1142-4_11

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1141-7

  • Online ISBN: 978-1-4939-1142-4

  • eBook Packages: Springer Protocols

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