Global Identification of O-GlcNAc-Modified Proteins

The O-linked N-acetylglucosamine (O-GlcNAc) modification of serine/threonine residues is an abundant posttranslational modification present in cytosolic and nuclear
proteins. The functions and subproteome of O-GlcNAc
modification remain largely undefined. Here we report the
application of the tagging-via-substrate (TAS) approach for
global identification of O-GlcNAc-modified proteins. The
TAS method utilizes an O-GlcNAc azide analogue for
metabolic labeling of O-GlcNAc-modified proteins, which
can be chemoselectively conjugated for detection and
enrichment of the proteins for proteomics studies. Our
study led to the identification of 199 putative O-GlcNAc-modified proteins from HeLa cells, among which 23 were
confirmed using reciprocal immunoprecipitation. Functional classification shows that proteins with diverse
functions are modified by O-GlcNAc, implying that O-GlcNAc might be involved in the regulation of multiple
cellular pathways.