Mechanism, Structure, and Inhibition of O-GlcNAc Processing Enzymes

Abstract

The post-translational modification of nucleocytoplasmic proteins with O-linked 2-acetamido-2-deoxy-D-glucopyranose (O-GlcNAc) is a topic of considerable interest and attracts a great deal of research effort. O-GlcNAcylation is a dynamic process which can occur multiple times over the lifetime of a protein, sometimes in a reciprocal relationship with phosphorylation. Several hundred proteins, which are involved in a diverse range of cellular processes, have been identified as being modified with the monosaccharide. The control of the O-GlcNAc modification state on different protein targets appears to be important in the aetiology of a number of diseases, including type II diabetes, neurodegenerative diseases and cancer. Two enzymes are responsible for the addition and removal of the O-GlcNAc modification: uridine diphospho-N-acetylglucosamine:polypeptide β-N-acetylglucosaminyltransferase (OGT) and O-GlcNAcase (OGA), respectively. Over the past decade the volume of information known about these two enzymes has increased significantly. In particular, mechanistic studies of OGA, in conjunction with structural studies of bacterial homologues of OGA have stimulated the design of inhibitors and offered a rationale for the binding of certain potent and selective inhibitors. Mechanistic information about OGT lags a little way behind OGA, but the recent deduction of the structure of an OGT bacterial homologue should now drive these studies forward.

Modulation of Epigenetic Targets for Anticancer Therapy: Clinicopathological Relevance, Structural Data and Drug Discovery Perspectives

“…In general, the addition of O-GlcNAc is reciprocal with Ser and Thr phosphorylation, either by modification of the same residue or nearby residues [520]. This PTM modification is regulated by only two enzymes: a glycosyltransferase that catalyzes the transfer of GlcNAc to substrate proteins, also known as O-GlcNAc transferase (OGT), and a glycoside hydrolase, also known as OGlcNAcase (OGA) or O-N-acetylglucosaminidase, that catalyzes the hydrolysis of the glycosidic linkage [521]. While it is interesting to note that in mammalians only these two highly conserved enzymes are responsible of O-GlcNAc cycling, it is worth emphasizing that the targeting of these enzymes is highly specific and is controlled by many interacting subunits.…”

“…Details about how OGT recognizes and glycosylates its protein substrates, including histone proteins, were almost unknown until recent years, when novel protein structural data became available (PDB codes: 1W3B and 3TAX) [521,534,535]. In 2011, the first two crystal structures of human OGT were solved: a binary complex with uridine 5'-diphosphate (UDP) and a ternary complex with UDP and a peptide substrate, including the catalytic region (PDB codes: 3PE3 and 3PE4) [536].…”

“…Nevertheless, small inhibitors for both enzymes, e.g. by PUGNAc and related derivatives, have been described in literature [521,[545][546][547][548]. It is expected that the discovery of other inhibitors of these enzymes, for use as cellular probes, will help the full understanding of OGlcNAc as a covalent histone modification and will contribute to foster research toward new epigenetic therapeutic agents.…”

O-Linked β-N-Acetylglucosamine (O-GlcNAc) Regulates Emerin Binding to Barrier to Autointegration Factor (BAF) in a Chromatin- and Lamin B-enriched “Niche”

“…Each reaction contained ϳ1 g of emerin polypeptide (wild type or mutated His-tagged emerin 1-220) plus 1 g of OGT and CIP and 1.0 Ci of [ 3 H]UDP-GlcNAc diluted with "cold" UDP-GlcNAc to a final GlcNAc concentration of 8.33 M (10-fold dilution of radiolabeled sugar). We chose this UDP-GlcNAc concentration to exceed the lowest reported K m for OGT (6,35, or 217 M) (73,74). Reactions were incubated overnight, quenched with SDS, resolved by SDS-PAGE, Coomassie-stained, and autoradiographed (Fig.…”

Alternating Phosphorylation with O-GlcNAc Modification: Another Way to Control Protein Function

“…For example, the short OGA exhibits comparative resistance to PugNAc and NAG-thiazoline, but is very sensitive to alpha-GlcNAc thiolsulfonate (Zachara & Hart, 2006). Inhibition of OGT and OGA represents an area of great interest on O-GlcNAcylation research, which is evident from the increasing number of studies addressing the enzymes molecular mechanisms for the addition and removal of O-GlcNAc (Borodkin & van Aalten, 2010;Dorfmueller et al, , 2011Gloster et al, 2011;Gloster & Vocadlo, 2010;Lameira et al, 2011;Lazarus et al, 2011;Li et al, 2011;Macauley & Vocadlo, 2010;MartinezFleites et al, 2010).…”