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- Title
A proteomics approach to understanding protein ubiquitination.
- Authors
Peng, Junmin; Schwartz, Daniel; Elias, Joshua E; Thoreen, Carson C; Cheng, Dongmei; Marsischky, Gerald; Roelofs, Jeroen; Finley, Daniel; Gygi, Steven P
- Abstract
There is a growing need for techniques that can identify and characterize protein modifications on a large or global scale. We report here a proteomics approach to enrich, recover, and identify ubiquitin conjugates from Saccharomyces cerevisiae lysate. Ubiquitin conjugates from a strain expressing 6xHis-tagged ubiquitin were isolated, proteolyzed with trypsin and analyzed by multidimensional liquid chromatography coupled with tandem mass spectrometry (LC/LC-MS/MS) for amino acid sequence determination. We identified 1,075 proteins from the sample. In addition, we detected 110 precise ubiquitination sites present in 72 ubiquitin-protein conjugates. Finally, ubiquitin itself was found to be modified at seven lysine residues providing evidence for unexpected diversity in polyubiquitin chain topology in vivo. The methodology described here provides a general tool for the large-scale analysis and characterization of protein ubiquitination.
- Publication
Nature biotechnology, 2003, Vol 21, Issue 8, p921
- ISSN
1087-0156
- Publication type
Journal Article
- DOI
10.1038/nbt849