Identification of SUMO-conjugated proteins and their SUMO attachment sites using proteomic mass spectrometry

Methods Mol Biol. 2009;497:33-49. doi: 10.1007/978-1-59745-566-4_3.

Abstract

The covalent modification of cellular factors by the small ubiquitin-like modifier (SUMO) has emerged as a key regulatory pathway for many biological processes. One recent advance in the field of SUMO modification that has provided important insights into SUMO-mediated regulatory networks is the ability to use proteomic mass spectrometry to identify the substrates of SUMO modification as well as their sites of conjugation (1-10). In this chapter, we describe a global strategy for affinity purifying and identifying a broad spectrum of SUMO-conjugated proteins and a focused approach for purifying a selected SUMO target and mapping its SUMO attachment site(s). Although both methods were initially developed for use in S. cerevisiae, they can be readily adapted to study the SUMO pathway in higher eukaryotes.

Publication types

  • Review

MeSH terms

  • Algorithms
  • Binding Sites
  • Mass Spectrometry / methods*
  • Models, Biological
  • Protein Processing, Post-Translational
  • Proteomics / methods*
  • SUMO-1 Protein / metabolism
  • Saccharomyces cerevisiae / chemistry
  • Saccharomyces cerevisiae / metabolism
  • Small Ubiquitin-Related Modifier Proteins / isolation & purification*
  • Small Ubiquitin-Related Modifier Proteins / metabolism*
  • Ubiquitin-Conjugating Enzymes / isolation & purification*
  • Ubiquitin-Conjugating Enzymes / metabolism

Substances

  • SUMO-1 Protein
  • Small Ubiquitin-Related Modifier Proteins
  • Ubiquitin-Conjugating Enzymes