Versatile O-GlcNAc transferase assay for high-throughput identification of enzyme variants, substrates, and inhibitors
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- Title
- Versatile O-GlcNAc transferase assay for high-throughput identification of enzyme variants, substrates, and inhibitors
- Author(s)
- E J Kim; L K Abramowitz; M R Bond; D C Love; D W Kang; H F Leucke; D W Kang; Jong Seog Ahn; J A Hanover
- Bibliographic Citation
- Bioconjugate Chemistry, vol. 25, no. 6, pp. 1025-1030
- Publication Year
- 2014
- Abstract
- The dynamic glycosylation of serine/threonine residues on nucleocytoplasmic proteins with a single N-acetylglucosamine (O-GlcNAcylation) is critical for many important cellular processes. Cellular O-GlcNAc levels are highly regulated by two enzymes: O-GlcNAc transferase (OGT) is responsible for GlcNAc addition and O-GlcNAcase (OGA) is responsible for removal of the sugar. The lack of a rapid and simple method for monitoring OGT activity has impeded the efficient discovery of potent OGT inhibitors. In this study we describe a novel, single-well OGT enzyme assay that utilizes 6 × His-tagged substrates, a chemoselective chemical reaction, and unpurified OGT. The high-throughput Ni-NTA Plate OGT Assay will facilitate discovery of potent OGT-specific inhibitors on versatile substrates and the characterization of new enzyme variants.
- ISSN
- 1043-1802
- Publisher
- Amer Chem Soc
- DOI
- http://dx.doi.org/10.1021/bc5001774
- Type
- Article
- Appears in Collections:
- Ochang Branch Institute > Chemical Biology Research Center > 1. Journal Articles
- Files in This Item:
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