Large-scale analysis of the human ubiquitin-related proteome

Proteomics. 2005 Nov;5(16):4145-51. doi: 10.1002/pmic.200401280.


Protein ubiquitylation contributes to the regulation of many cellular processes including protein degradation, receptor internalization, and repair of DNA damage. We now present a comprehensive characterization of ubiquitin-conjugated and ubiquitin-associated proteins in human cells. The proteins were purified by immunoaffinity chromatography under denaturing or native conditions. They were then digested with trypsin, and the resulting peptides were analyzed by 2-D LC and MS/MS. A total of 670 distinct proteins were identified; 345 proteins (51%) were classified as Urp-D (ubiquitin-related proteome under the denaturing condition) and comprised ubiquitin-conjugated molecules, whereas 325 proteins (49%) were classified as Urp-N (ubiquitin-related proteome only under the native condition) and included molecules that associated with ubiquitylated proteins. The proportions of proteins in various functional categories differed substantially between Urp-D and Urp-N. Many ribosomal subunits were detected in the Urp-D group of proteins and several of these subunits were directly shown to be ubiquitylated by mass spectrometric analysis, suggesting that ubiquitylation might play an important role in the regulation and/or quality control of ribosomal proteins. Our results demonstrate the potential of proteomics analysis of protein ubiquitylation to provide important insight into the regulation of protein stability and other ubiquitin-related cellular functions.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • Cell Line
  • Chromatography, Affinity
  • Chromatography, Liquid
  • Humans
  • Mass Spectrometry
  • Protein Binding
  • Protein Subunits / chemistry
  • Protein Subunits / metabolism
  • Proteome / analysis*
  • Proteome / metabolism
  • Ribosomes / chemistry
  • Ribosomes / metabolism
  • Ubiquitin / analysis*
  • Ubiquitin / metabolism


  • Protein Subunits
  • Proteome
  • Ubiquitin