Detection and identification of O -GlcNAc-modified proteins using 6-azido-6-deoxy- N -acetyl-galactosamine

• Published in: Organic & biomolecular chemistry Vol. 17; no. 17; pp. 4326 - 4334
• Main Authors: Guo, Jianshuang, Zhang, Guoqiang, Ma, Jing, Zhao, Caili, Xue, Qingqing, Wang, Jiyan, Liu, Wenjie, Liu, Kaihui, Wang, Haifeng, Liu, Ning, Song, Qitao, Li, Jing
• Format: Journal Article
• Language: English
• Published: England Royal Society of Chemistry 24.04.2019
• Subjects: Animals; beta-N-Acetylhexosaminidases - analysis; beta-N-Acetylhexosaminidases - metabolism; Cell lines; Cells, Cultured; Chemical synthesis; D-Galactosamine; Galactosamine - analogs & derivatives; Galactosamine - chemical synthesis; Galactosamine - chemistry; Glycosylation; Humans; Lysates; Mice; Molecular Probes - chemical synthesis; Molecular Probes - chemistry; Molecular Structure; Monosaccharides; N-Acetylgalactosamine; N-Acetylglucosaminyltransferases - analysis; N-Acetylglucosaminyltransferases - metabolism; Organic chemistry; Proteins; Proteomics
• ISSN: 1477-0520; 1477-0539; 1477-0539
• DOI: 10.1039/C9OB00516A

An unnatural monosaccharide with a C6-azide, Ac 3 6AzGalNAc, has been developed as a potent and selective probe for O -GlcNAc-modified proteins. Combined with click chemistry, we demonstrate that Ac 3 6AzGalNAc can robustly label O -GlcNAc glycosylation in a wide range of cell lines. Meanwhile, cell imaging and LC-MS/MS proteomics verify its selective activity on O -GlcNAc. More importantly, the protocol presented here provides a general methodology for tracking, capturing and identifying unnatural monosaccharide modified proteins in cells or cell lysates.