Inhibition of recombinant human carboxylesterase 1 and 2 and monoacylglycerol lipase by chlorpyrifos oxon, paraoxon and methyl paraoxon

Abstract

Oxons are the bioactivated metabolites of organophosphorus insecticides formed via cytochrome P450 monooxygenase-catalyzed desulfuration of the parent compound. Oxons react covalently with the active site serine residue of serine hydrolases, thereby inactivating the enzyme. A number of serine hydrolases other than acetylcholinesterase, the canonical target of oxons, have been reported to react with and be inhibited by oxons. These off-target serine hydrolases include carboxylesterase 1 (CES1), CES2, and monoacylglycerol lipase. Carboxylesterases (CES, EC 3.1.1.1) metabolize a number of xenobiotic and endobiotic compounds containing ester, amide, and thioester bonds and are important in the metabolism of many pharmaceuticals. Monoglyceride lipase (MGL, EC 3.1.1.23) hydrolyzes monoglycerides including the endocannabinoid, 2-arachidonoylglycerol (2-AG). The physiological consequences and toxicity related to the inhibition of off-target serine hydrolases by oxons due to chronic, low level environmental exposures are poorly understood. Here, we determined the potency of inhibition (IC 50 values; 15 min preincubation, enzyme and inhibitor) of recombinant CES1, CES2, and MGL by chlorpyrifos oxon, paraoxon and methyl paraoxon. The order of potency for these three oxons with CES1, CES2, and MGL was chlorpyrifos oxon > paraoxon > methyl paraoxon, although the difference in potency for chlorpyrifos oxon with CES1 and CES2 did not reach statistical significance. We also determined the bimolecular rate constants (k inact /K I ) for the covalent reaction of chlorpyrifos oxon, paraoxon and methyl paraoxon with CES1 and CES2. Consistent with the results for the IC 50 values, the order of reactivity for each of the three oxons with CES1 and CES2 was chlorpyrifos oxon > paraoxon > methyl paraoxon. The bimolecular rate constant for the reaction of chlorpyrifos oxon with MGL was also determined and was less than the values determined for chlorpyrifos oxon with CES1 and CES2 respectively. Together, the results define the kinetics of inhibition of three important hydrolytic enzymes by activated metabolites of widely used agrochemicals.

Identification of the Major Prostaglandin Glycerol Ester Hydrolase in Human Cancer Cells

“…Consistent with the wide substrate acceptance exhibited by CES1 and PPT1, both enzymes are capable of hydrolyzing PG-Gs and 2-AG (24,30,31). In THP1 monocytes, the hydrolysis of 2-AG is almost entirely attributed to CES1, with minor involvement of PPT1 (24,30,31). Kinetic analysis of both enzymes showed almost 2-fold greater catalytic turnover for 2-AG than for PG-Gs (31).…”

“…Similarly, PPT1, a lysosomal hydrolase, has multiple substrates; however, it is predominantly responsible for the depalmitoylation of a number of proteins as well as hydrolysis of palmitoyl-CoA and palmitoyl thioglucoside (28,29). Consistent with the wide substrate acceptance exhibited by CES1 and PPT1, both enzymes are capable of hydrolyzing PG-Gs and 2-AG (24,30,31). In THP1 monocytes, the hydrolysis of 2-AG is almost entirely attributed to CES1, with minor involvement of PPT1 (24,30,31).…”

Specific Inter-residue Interactions as Determinants of Human Monoacylglycerol Lipase Catalytic Competency

“…Ligand binding experiments were carried out using a 50 mM stock solution of paraoxon (diethyl 4-nitrophenyl phosphate), dissolved in DMSO-d 6 (18). Typically, no more than 5 l of paraoxon solution was added to each enzyme sample to achieve a concentration of paraoxon twice that of the enzyme.…”

Increased cancer burden among pesticide applicators and others due to pesticide exposure

“…Oxons are exquisitely potent compounds that inhibit serine hydrolases via covalent modification of the catalytic serine residue in the enzyme active site. 104 Serine hydrolases participate in a wide variety of physiological and pathophysiological processes, including signal transduction in neural tissue, digestion, immune response, xenobiotic detoxification, and the clotting cascade. Thus, inhibition of these enzymes may lead to a variety of pathological effects.…”