Analysis of ubiquitinated proteome by quantitative mass spectrometry

Chan Hyun Na, Junmin Peng

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Protein modification by ubiquitin (Ub) is one of the most common posttranslational events in eukaryotic cells. Ubiquitinated proteins are destined to various fates such as proteasomal degradation, protein trafficking, DNA repair, and immune response. In the last decade, vast improvements of mass spectrometry make it feasible to analyze the minute amount of ubiquitinated components in vivo. When combined with quantitative strategies, such as stable isotope labeling with amino acids in cell culture (SILAC), it is capable of profiling ubiquitinated proteome under different experimental conditions. Here, we describe a procedure to perform such a study, including differential protein labeling by the SILAC method, enrichment of ubiquitinated species, mass spectrometric analysis, and quality control to reduce false positives. The potential challenges and limitations of the procedure are also discussed.

Original languageEnglish (US)
Title of host publicationQuantitative Methods in Proteomics
EditorsMarcus Katrin
Pages417-429
Number of pages13
DOIs
StatePublished - 2012

Publication series

NameMethods in Molecular Biology
Volume893
ISSN (Print)1064-3745

Keywords

  • Mass spectrometry
  • Proteomics
  • SILAC
  • Ubiquitin

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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