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Updated: Apr 29, 2026
High-throughput Screening of Carbohydrate-degrading Enzymes Using Novel Insoluble Chromogenic Substrate Assay Kits
Published on: September 20, 2016
Versatile O-GlcNAc transferase assay for high-throughput identification of enzyme variants, substrates, and
Eun J Kim1, Lara K Abramowitz, Michelle R Bond
1Department of Science Education-Chemistry Major, Daegu University , Gyeongbuk 712-714, South Korea.
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.
