Design of inhibitors of orotidine monophosphate decarboxylase using bioisosteric replacement and determination of inhibition kinetics.
Inhibitors of orotidine monophosphate decarboxylase (ODCase) have applications in RNA viral, parasitic, and other infectious diseases. ODCase catalyzes the decarboxylation of orotidine monophosphate (OMP), producing uridine monophosphate (UMP). Novel inhibitors 6-amino-UMP and 6-cyano-UMP were designed on the basis of the substructure volumes in the substrate OMP and in an inhibitor of ODCase, barbituric acid monophosphate, BMP. A new enzyme assay method using isothermal titration calorimetry (ITC) was developed to investigate the inhibition kinetics of ODCase. The reaction rates were measured by monitoring the heat generated during the decarboxylation reaction of orotidine monophosphate. Kinetic parameters (k(cat) = 21 s(-1) and KM = 5 microM) and the molar enthalpy (DeltaH(app) = 5 kcal/mol) were determined for the decarboxylation of the substrate by ODCase. Competitive inhibition of the enzyme was observed and the inhibition constants (Ki) were determined to be 12.4 microM and 29 microM for 6-aza-UMP and 6-cyano-UMP, respectively. 6-Amino-UMP was found to be among the potent inhibitors of ODCase, having an inhibition constant of 840 nM. We reveal here the first inhibitors of ODCase designed by the principles of bioisosterism and a novel method of using isothermal calorimetry for enzyme inhibition studies.
Poduch
Ewa
E
Molecular Design and Information Technology Center, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario M5S 2S2, Canada.
Bello
Angelica M
AM
Tang
Sishi
S
Fujihashi
Masahiro
M
Pai
Emil F
EF
Kotra
Lakshmi P
LP
D002151:Calorimetry; D003198:Computer Simulation; D015195:Drug Design; D007700:Kinetics; D008958:Models, Molecular; D009964:Orotidine-5'-Phosphate Decarboxylase / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D013816:Thermodynamics; D014542:Uridine Monophosphate / Q000031:analogs & derivatives* / Q000138:chemical synthesis* / Q000737:chemistry
C513368:6-aminouridine 5'-O-monophosphate; C513367:6-cyanouridine 5'-O-monophosphate; C011569:6-azauridine-5'-monophosphate; D014542:Uridine Monophosphate; D009964:Orotidine-5'-Phosphate Decarboxylase