Mass spectrometric determination of protein ubiquitination.

Carol E. Parker, Viorel Mocanu, Maria R. Warren, Susanna F. Greer, Christoph H. Borchers

Research output: Contribution to journalArticlepeer-review

9 Citations (Scopus)

Abstract

Mass spectrometric methods of determining protein ubiquitination are described. Characteristic mass shifts and fragment ions indicating ubiquitinated lysine residues in tryptic and gluC digests are discussed. When a ubiquitinated protein is enzymatically digested, a portion of the ubiquitin side chain remains attached to the modified lysine. The ubiquitinated peptide thus has two N-termini- one from the original peptide and one from the ubiquitin side chain. Thus, it is possible to have two series of b ions and y ions, the additional series is the one that includes fragments containing portions of the ubiquitin side chain. Any diagnostic ions for the modification must include portions of this side chain. Fragment ions involving any part of the "normal" peptide will vary in mass according to the peptide being modified and will therefore not be of general diagnostic use. These diagnostic ions, found through examination of the MS/MS spectra of model ubiquitinated tryptic and gluC peptides, can be used to trigger precursor ion scanning in automated MS/MS data acquisition scanning modes.

Original languageEnglish
Pages (from-to)153-173
Number of pages21
JournalMethods in Molecular Biology
Volume301
DOIs
Publication statusPublished - 2005
Externally publishedYes

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