Figure 2

Scheme for the enrichment of O -GlcNAcylated proteins/peptides. Most commonly used strategies with antibody enrichment (1), lectin enrichment (2), BEMAD (3), chemoenzymatic labeling (4) and metabolic labeling (5) are illustrated. In (1), proteins are captured onto antibody/antibodies-conjugated beads, and the enriched ones are digested and identified by tandem mass spectrometry. In (2), (3), and (4), proteins are digested into peptides, which are captured with lectin-conjugated resin (2), thio-capture column after BEMAD (3), and streptavidin-conjugated beads after chemoenzymatic labeling (4), with the enriched peptides identified by tandem mass spectrometry. In (5), cells are fed with GlcNAc analogs GlcNAz and GlcNAlk, and the GlcNAz- and GlcNAlk-containing proteins are subjected to click chemistry, streptavidin-conjugated beads enrichment and digestion, with the digests analyzed by tandem mass spectrometry. Note: The cocktail usage of several methods (e.g., chemo-enzymatic/metabolic labeling and BEMAD) has also been applied in some cases.