Discovery of O-GlcNAc-modified proteins in published large-scale proteome data.
The attachment of N-acetylglucosamine to serine or threonine residues (O-GlcNAc) is a post-translational modification on nuclear and cytoplasmic proteins with emerging roles in numerous cellular processes, such as signal transduction, transcription, and translation. It is further presumed that O-GlcNAc can exhibit a site-specific, dynamic and possibly functional interplay with phosphorylation. O-GlcNAc proteins are commonly identified by tandem mass spectrometry following some form of biochemical enrichment. In the present study, we assessed if, and to which extent, O-GlcNAc-modified proteins can be discovered from existing large-scale proteome data sets. To this end, we conceived a straightforward O-GlcNAc identification strategy based on our recently developed Oscore software that automatically analyzes tandem mass spectra for the presence and intensity of O-GlcNAc diagnostic fragment ions. Using the Oscore, we discovered hundreds of O-GlcNAc peptides not initially identified in these studies, and most of which have not been described before. Merely re-searching this data extended the number of known O-GlcNAc proteins by almost 100 suggesting that this modification exists even more widely than previously anticipated and the modification is often sufficiently abundant to be detected without enrichment. However, a comparison of O-GlcNAc and phospho-identifications from the very same data indicates that the O-GlcNAc modification is considerably less abundant than phosphorylation. The discovery of numerous doubly modified peptides (i.e. peptides with one or multiple O-GlcNAc or phosphate moieties), suggests that O-GlcNAc and phosphorylation are not necessarily mutually exclusive, but can occur simultaneously at adjacent sites.
Center of Life and Food Sciences Weihenstephan, Technische Universität München, Emil-Erlenmeyer-Forum 5, 85354 Freising, Germany.
Hannes
H
Hahne
Amin Moghaddas
AM
Gholami
Bernhard
B
Kuster
D000117:Acetylglucosamine / Q000378:metabolism*; D000595:Amino Acid Sequence; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D030562:Databases, Protein; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis*; D010750:Phosphoproteins / Q000032:analysis*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D020543:Proteome / Q000032:analysis / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D012984:Software; D053719:Tandem Mass Spectrometry; D013912:Threonine / Q000378:metabolism
D010455:Peptides; D010750:Phosphoproteins; D020543:Proteome; D013912:Threonine; D012694:Serine; D000117:Acetylglucosamine