Kinetics of substrate hydrolysis and inhibition by mipafox of paraoxon-preinhibited hen brain esterase activity.
Biochemical J 1986 Jun; 236(2):503-507
The kinetics of substrate hydrolysis and the kinetics of inhibition by mipafox (371868) of the paraoxon (311455) resistant esterases in a membrane fraction of White Leghorn hen brain were investigated. Paraoxon was added to homogenized hen brain to a concentration of 40 micromoles and the homogenate was processed with a series of centrifugation procedures. Paraoxon preinhibited hen brain was found to have both neurotoxicant sensitive and insensitive esterase components. Kinetic parameters for the hydrolysis of phenyl- valerate (PV) and phenyl-phenylacetate (PPA) were as follows: neurotoxic esterase (NTE) Michaelis constant (Km) values were found to be about 0.0014 and 0.00016 Molar (M), respectively, at 37 degrees-C and 0.0024M for PV at 25 degrees-C. NTE kinetic constants for mipafox with either PV or PPA as substrate were determined to be Km=0.00018M and the phosphorylation constant (K2)=1.1min(-1) at 37 degrees-C; at 25 degrees-C, Km=0.0006M and K2=0.7min(-1) for PV only. The 25 degree-C data was analyzed with a two component model with or without formation of Michaelis complex on NTE, or a three component model without formation of Michaelis complex. The authors report that the Michaelis model fit the data significantly better and conclude that the higher apparent Ki values at low mipafox concentrations are due to formation of Michaelis complex at high concentrations. The authors also conclude that if mipafox competes directly for the active site of NTE, then the much greater inhibitory power of mipafox must be due to a much greater K2 for mipafox than for paraoxon.
NIOSH-Grant; NIOSH-Publication; Phenols; Organophosphorous-compounds; Nitrophenols; Fluorinated-hydrocarbons; Halogenated-hydrocarbons; Halogenated-phosphates; Laboratory-animals; Esters; Enzyme-activity; Enzyme-inhibitors; Neurotoxicity
Mohamed B. Abou-Donia, Department of Pharmacology, Duke University Medical Center, Durham, NC 27710, U.S.A.
Pharmacology Duke University Medical CenterBox 3813 Durham, NC 27710