Visualization of O-GlcNAc glycosylation stoichiometry and dynamics using resolvable poly(ethylene glycol) mass tags

Curr Protoc Chem Biol. 2013;5(4):281-302. doi: 10.1002/9780470559277.ch130153.


O-linked N-acetylglucosamine (O-GlcNAc) glycosylation is a dynamic protein posttranslational modification with roles in processes such as transcription, cell cycle regulation, and metabolism. Detailed mechanistic studies of O-GlcNAc have been hindered by a lack of methods for measuring O-GlcNAc stoichiometries and the interplay of glycosylation with other posttranslational modifications. We recently developed a method for labeling O-GlcNAc-modified proteins with resolvable poly(ethylene glycol) mass tags. This mass-tagging approach enables the direct measurement of glycosylation stoichiometries and the visualization of distinct O-GlcNAc-modified subpopulations. Here, we describe procedures for labeling O-GlcNAc glycoproteins in cell lysates with mass tags.

Keywords: O-linked N-acetylglucosamine; chemoenzymatic labeling; glycosylation; poly(ethylene glycol); posttranslational modifications; protein subpopulations.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Acetylglucosamine / metabolism*
  • Animals
  • Azides / chemistry
  • Carbohydrate Sequence
  • Cycloaddition Reaction / methods
  • Galactosyltransferases / chemistry
  • Galactosyltransferases / genetics
  • Glycosylation / drug effects*
  • Humans
  • Molecular Sequence Data
  • Oximes / metabolism
  • Polyethylene Glycols / pharmacology*
  • Protein Processing, Post-Translational / genetics
  • Proteins / chemistry


  • Azides
  • Oximes
  • Proteins
  • Polyethylene Glycols
  • Galactosyltransferases
  • Acetylglucosamine