Since the discovery of O-GlcNAc modification (O-GlcNAcylation) 20 years ago, much attention has been given to OGT (O-GlcNAc transferase), the unique enzyme responsible for the nuclear and cytosolic O-GlcNAcylation processes. This review focuses on protocols that are routinely used to analyze OGT expression and activity. First are detailed techniques using rabbit polyclonal anti-OGT antibodies, namely, Western blot, (co-)immunoprecipitation, and immunofluorescence. We also describe the measurement of OGT activity by using synthetic peptides as acceptors and radiolabeled UDP-GlcNAc. Finally, a sensitive HPAEC-based technique to measure the cellular content of UDP-GlcNAc, the donor substrate of OGT, is described in detail. © Springer Science+Business Media New York 2013.
CITATION STYLE
Lefebvre, T., Drougat, L., Stichelen, S. O. V., Michalski, J. C., & Vercoutter-Edouart, A. S. (2013). Antibodies and activity measurements for the detection of O-GlcNAc transferase and assay of its substrate, UDP-GlcNAc. Methods in Molecular Biology, 1022, 147–159. https://doi.org/10.1007/978-1-62703-465-4_12
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