Advances in chemical probing of protein -GlcNAc glycosylation: structural role and molecular mechanisms.

The addition of -linked-β-D--acetylglucosamine (-GlcNAc) onto serine and threonine residues of nuclear and cytoplasmic proteins is an abundant, unique post-translational modification governing important biological processes. -GlcNAc dysregulation underlies several metabolic disorders leading to human diseases, including cancer, neurodegeneration and diabetes. This review provides an extensive summary of the recent progress in probing -GlcNAcylation using mainly chemical methods, with a special focus on discussing mechanistic insights and the structural role of -GlcNAc at the molecular level. We highlight key aspects of the -GlcNAc enzymes, including development of OGT and OGA small-molecule inhibitors, and describe a variety of chemoenzymatic and chemical biology approaches for the study of -GlcNAcylation. Special emphasis is placed on the power of chemistry in the form of synthetic glycopeptide and glycoprotein tools for investigating the site-specific functional consequences of the modification. Finally, we discuss in detail the conformational effects of -GlcNAc glycosylation on protein structure and stability, relevant -GlcNAc-mediated protein interactions and its molecular recognition features by biological receptors. Future research in this field will provide novel, more effective chemical strategies and probes for the molecular interrogation of -GlcNAcylation, elucidating new mechanisms and functional roles of -GlcNAc with potential therapeutic applications in human health.