Ultradeep O-GlcNAc proteomics reveals widespread O-GlcNAcylation on tyrosine residues of proteins
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ABSTRACT: We developed an ultra-deep O-GlcNAc proteomics workflow, by integrating multiple-proteases digestion, two mass spectrometric approaches (i.e., EThcD fragmentation and HCD product-dependent EThcD fragmentation), and two data analysis tools (i.e., MaxQuant and Proteome Discoverer). The performance of this strategy was benchmarked by the analysis of whole lysates from PANC-1 (a pancreatic cancer cell line), in total 2831 O-GlcNAc sites were unambiguously identified, representing the largest O-GlcNAc dataset of an individual study reported so far. Unexpectedly, in addition to confirming known sites and discovering many novel sites of Ser/Thr modification, O-GlcNAcylation was found on 121 tyrosine (Tyr) residues of 93 proteins. Through in vitro enzymatic assays, we revealed that OGT shows catalytic capacity to transfer O-GlcNAc onto Tyr residues of peptides and OGA can remove Tyr O-GlcNAcylation from peptides. Taken together, we discovered widespread O-GlcNAcylation on tyrosine residues of proteins and Tyr O-GlcNAcylation is mediated by OGT and OGA. As a novel form of glycosylation, Tyr O-GlcNAcylation may have important regulatory roles.
INSTRUMENT(S): Orbitrap Fusion Lumos
ORGANISM(S): Homo Sapiens (ncbitaxon:9606)
SUBMITTER: Junfeng Ma
PROVIDER: MSV000094669 | MassIVE | Fri May 03 08:23:00 BST 2024
REPOSITORIES: MassIVE
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