nitrophenols and methyl-demeton

Glycerophosphodiesterase (GpdQ) from Enterobacter aerogenes is a nonspecific diesterase that enables Escherichia coli to utilize alkyl phosphodiesters, such as diethyl phosphate, as the sole phosphorus source. The catalytic properties of GpdQ were determined, and the best substrate found was bis(p-nitrophenyl) phosphate with a kcat/Km value of 6.7 x 10(3) M-1 s-1. In addition, the E. aerogenes diesterase was tested as a catalyst for the hydrolysis of a series of phosphonate monoesters which are the hydrolysis products of the highly toxic organophosphonate nerve agents sarin, soman, GF, VX, and rVX. Among the phosphonate monoesters tested, the hydrolysis product of rVX, isobutyl methyl phosphonate, was the best substrate with a kcat/Km value of 33 M-1 s-1. The ability of GpdQ to hydrolyze the phosphonate monoesters provides an alternative selection strategy in the search of enhanced variants of the bacterial phosphotriesterase (PTE) for the hydrolysis of organophosphonate nerve agents. This investigation demonstrated that the previously reported activity of GpdQ toward the hydrolysis of methyl demeton-S is due to the presence of a diester contaminant in the commercial material. Furthermore, it was shown that GpdQ is capable of hydrolyzing a close analogue of EA 2192, the most toxic and persistent degradation product of the nerve agent VX.

Topics: Chemical Warfare Agents; Cholinesterase Inhibitors; Disulfoton; Enterobacter aerogenes; Kinetics; Microbial Viability; Molecular Structure; Mutation; Nitrophenols; Organophosphates; Organophosphonates; Organothiophosphorus Compounds; Paraoxon; Phenols; Phosphoric Diester Hydrolases; Phosphoric Triester Hydrolases; Recombinant Proteins; Substrate Specificity

1. The kinetics and inhibition of p-nitrophenylacetate hydrolysis by cytosolic esterases of female alfalfa leafcutting bees, Megachile rotundata (Fab.) was examined. 2. For p-nitrophenylacetate, the Km = 1.24 x 10(-4) M and Vmax = 2.29 x 10(-9) mol/s per mg protein. 3. Regarding four organophosphate insecticides, the mechanism of inhibition in all cases was mixed (competitive and uncompetitive) and, based on inhibition constants, the order of toxicity was naled greater than paraoxon greater than trichlorfon greater than oxydemeton methyl. 4. Comparisons are made to the honey bee, Apis mellifera.

Topics: Animals; Bees; Esterases; Female; Hydrolysis; Insecticides; Kinetics; Naled; Nitrophenols; Organothiophosphorus Compounds; Paraoxon; Trichlorfon