GlcNAcstatins are nanomolar inhibitors of human O-GlcNAcase inducing cellular hyper-O-GlcNAcylation
Models, Molecular
0301 basic medicine
570
GlcNAcstatin
Glycosylation
Acylation
Blotting, Western
Catalysis
Acetylglucosamine
03 medical and health sciences
Bacterial Proteins
In vivo
Humans
Nanotechnology
beta-D-glucosaminidase
Enzyme Inhibitors
GlcNAc modification
Inhibition
0303 health sciences
Linked-N-acetylglucosamine
Dose-Response Relationship, Drug
Molecular Structure
Proteins
Life Sciences
Insulin resistance
O-GlcNAcase
Alzheimer's disease
beta-N-Acetylhexosaminidases
Kinetics
Mutagenesis
O-GlcNAc
Structural insights
Mechanism
Research Article
Protein Binding
DOI:
10.1042/bj20090110
Publication Date:
2009-03-17T15:32:00Z
AUTHORS (4)
ABSTRACT
O-GlcNAcylation is an essential, dynamic and inducible post-translational glycosylation of cytosolic proteins in metazoa and can show interplay with protein phosphorylation. Inhibition of OGA (O-GlcNAcase), the enzyme that removes O-GlcNAc from O-GlcNAcylated proteins, is a useful strategy to probe the role of this modification in a range of cellular processes. In the present study, we report the rational design and evaluation of GlcNAcstatins, a family of potent, competitive and selective inhibitors of human OGA. Kinetic experiments with recombinant human OGA reveal that the GlcNAcstatins are the most potent human OGA inhibitors reported to date, inhibiting the enzyme in the sub-nanomolar to nanomolar range. Modification of the GlcNAcstatin N-acetyl group leads to up to 160-fold selectivity against the human lysosomal hexosaminidases which employ a similar substrate-assisted catalytic mechanism. Mutagenesis studies in a bacterial OGA, guided by the structure of a GlcNAcstatin complex, provides insight into the role of conserved residues in the human OGA active site. GlcNAcstatins are cell-permeant and, at low nanomolar concentrations, effectively modulate intracellular O-GlcNAc levels through inhibition of OGA, in a range of human cell lines. Thus these compounds are potent selective tools to study the cell biology of O-GlcNAc.
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