Inhibition of a bacterial O-GlcNAcase homologue by lactone and lactam derivatives: structural, kinetic and thermodynamic analyses.
The dynamic, intracellular, O-GlcNAc modification is of continuing interest and one whose study through targeted "chemical genetics" approaches is set to increase. Of particular importance is the inhibition of the O-GlcNAc hydrolase, O-GlcNAcase (OGA), since this provides a route to elevate cellular O-GlcNAc levels, and subsequent phenotypic evaluation. Such a small molecule approach complements other methods and potentially avoids changes in protein-protein interactions that manifest themselves in molecular biological approaches to O-GlcNAc transferase knockout or over-expression. Here we describe the kinetic, thermodynamic and three-dimensional structural analysis of a bacterial OGA analogue from Bacteroides thetaiotaomicron, BtGH84, in complex with a lactone oxime (LOGNAc) and a lactam form of N-acetylglucosamine and compare their binding signatures with that of the more potent inhibitor O-(2-acetamido-2-deoxy-D-glucopyranosylidene)amino N-phenyl carbamate (PUGNAc). We show that both LOGNAc and the N-acetyl gluconolactam are significantly poorer inhibitors than PUGNAc, which may reflect poorer mimicry of transition state geometry and steric clashes with the enzyme upon binding; drawbacks that the phenyl carbamate adornment of PUGNAc helps mitigate. Implications for the design of future generations of inhibitors are discussed.
He
Yuan
Y
York Structural Biology Laboratory, Department of Chemistry, The University of York, York, UK.
Bubb
Abigail K
AK
Stubbs
Keith A
KA
Gloster
Tracey M
TM
Davies
Gideon J
GJ
D001426:Bacterial Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001439:Bacteroides / Q000737:chemistry / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000494:pharmacology; D007700:Kinetics; D007769:Lactams / Q000737:chemistry* / Q000494:pharmacology; D007783:Lactones / Q000737:chemistry* / Q000494:pharmacology; D008958:Models, Molecular; D017351:N-Acetylglucosaminyltransferases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D013816:Thermodynamics
D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D007769:Lactams; D007783:Lactones; D017351:N-Acetylglucosaminyltransferases; C449880:O-GlcNAc transferase