Two independent research teams have zeroed in on the workings of O-linked β-N-acetylglucosamine transferase (OGT), an enzyme that adds the sugar N-acetylglucosamine (GlcNAc) selectively to various proteins. Although the groups come to different conclusions about a key detail of the mechanism, the work suggests that confirmation of the enzyme’s long-sought mode of action may be near.
Scientists are interested in how OGT works because the sugars it appends to proteins affect biological processes like embryo formation, gene transcription, responses to stress, and sensing of nutrients. Errors of OGT glycosylation have been associated with diabetes, cancer, and neurodegenerative diseases. But the way OGT catalyzes glycosylation is not yet well understood.
Now, a group led by Daan M. F. van Aalten of the University of Dundee, in Scotland, and another headed by Suzanne Walker of Harvard Medical School have obtained complexes of OGT with substrates, substrate analogs, and products and used crystallography, chemical probes, and kinetics to understand exactly how the complexes glycosylate proteins (Nat. Chem. Biol., DOI: 10.1038/nchembio.1108 and 10.1038/nchembio.1109).
The findings have important implications for ongoing efforts to pharmacologically interfere with the glycosylation process, comments John A. Hanover of NIH’s National Institute of Diabetes & Digestive & Kidney Diseases, an expert on O-linked GlcNAc.
OGT’s action requires proton abstraction from the hydroxyl of serine, one of two amino acids OGT can glycosylate. But the studies differ as to the catalytic base that carries this out. Walker’s team proposes that a water molecule abstracts serine’s proton in a process that may involve protonation of a nearby aspartate, whereas van Aalten’s group proposes that the activated donor sugar’s diphosphate abstracts the proton. Either way, the serine oxygen then links up with a carbon on the sugar ring. This expels uridine diphosphate from the activated donor substrate and completes the glycosylation process. The difference regarding the abstraction step will hopefully be resolved by future studies.
Hanover notes that both groups come to the surprising conclusion that the catalytic base does not come from the enzyme itself, as had been speculated. “The catalytic models proposed are not mutually exclusive,” he notes. “All of the features described are likely to play a key role in catalysis.”
The new work builds on other recent work on OGT: A Harvard Medical School team that included Walker, Michael B. Lazarus, and Piotr Sliz obtained the first high-resolution crystal structures of the enzyme; David J. Vocadlo and coworkers at Simon Fraser University, in Burnaby, British Columbia, identified a substrate analog that slows the enzyme’s action, making it easier to study; and the Walker lab showed that uridine diphosphate-GlcNAc, the activated form of the donor sugar, binds to the enzyme before glycosylation occurs.