Esterases acetylcholinesterase inhibition

Pesticide assessment guidelines under the Federal Insecticide, Fungicide, and Rodenticide Act stipulate that organophosphates proposed for use as insecticides be tested both for their capability to cause acute toxicities due to inhibition of acetylcholinesterase and for their potential to cause inhibition of neurotoxic esterase and subsequent delayed neuropathy. Testing could be performed in laboratory rodents because they, like all species, are susceptible to acetylcholinesterase inhibition, but rodents do not develop notable ataxia, and neuropathological... 

Sogorb, M.A., Gonz lez-Gonz lez, L, Pamies, D., et al., 2010. An alternative in vitro method for detecting neuropathic compounds based on acetylcholinesterase inhibition and on inhibition and aging of neuropathy target esterase (NTE). Toxicol. In Vitro 24 (3), 942-952. 

As discussed above, OPs are enzyme inhibitors, most obviously antiesterases and the most important esterase OPs inhibit is acetylcholinesterase. However, there is increasing evidence that some toxic effects are not mediated by inhibition of acetylcholinesterase. In fact, OPs bind to a variety of enzymes including esterases other than acetylcholinesterase e.g. carboxylesterase, long chain fatty acid hydrolase), serine peptidases, amidases and proteases and others, often modulating them (see review by Lockridge and Schopfer and section 10.3.2.1). ... 

This process of aging is believed to be critical in the development of delayed neuropathy, after NTE has been phosphorylated by an OP (see Chapter 10, Section 10.2.4). It is believed that most, if not all, of the B-esterases are sensitive to inhibition by OPs because they, too, have reactive serine at their active sites. It is important to emphasize that the interaction shown in Fignre 2.11 occurs with OPs that contain an oxon group. Phosphorothionates, which contain instead a thion group, do not readily interact in this way. Many OP insecticides are phosphorothionates, but these need to be converted to phosphate (oxon) forms by oxidative desulfuration before inhibition of acetylcholinesterase can proceed to any significant extent (see Section 2.3.2.2). 

The development of nerve gases in World War II, especially di-isopropylphosphofluoridate, (DIPF), promoted urgent investigations of their bases of action. Adrian et al. and Mackworth showed esterases, particularly acetylcholinesterase, were strongly inhibited (1941). The range of hydrolases sensitive to DIPF was extensive and included chymotrypsin and trypsin, both of which were available in purified crystalline form. DI PF was used to inhibit either enzyme, after which... 

Metabolites that are less reactive than suicide inhibitors may impact more distant enzymes, within the same cell, adjacent cells, or even in other tissues and organs, far removed from the original site of primary metabolism. For example, organopho-sphates (OPs), an ingredient in many pesticides, are metabolized by hepatic CYPs to intermediates, which, when transported to the nervous system, inhibit esterases that are critical for neural function. Acetylcholinesterase (AChE) catalyzes the hydrolysis of the ester bond in the neurotransmitter, acetylcholine, allowing choline to be recycled by the presynaptic neurons. If AChE is not effectively hydrolyzed by AChE in this manner, it builds up in the synapse and causes hyperexcitation of the postsynaptic receptors. The metabolites of certain insecticides, such as the phos-phorothionates (e.g., parathion and malathion) inhibit AChE-mediated hydrolysis. Phosphorothionates contain a sulfur atom that is double-bonded to the central phosphorus. However, in a CYP-catalyzed desulfuration reaction, the S atom is... 

Esterase activity is important in both the detoxication of organophosphates and the toxicity caused by them. Thus brain acetylcholinesterase is inhibited by organophosphates such as paraoxon and malaoxon, their oxidized metabolites (see above). This leads to toxic effects. Malathion, a widely used insecticide, is metabolized mostly by carboxylesterase in mammals, and this is a route of detoxication. However, an isomer, isomalathion, formed from malathion when solutions are inappropriately stored, is a potent inhibitor of the carboxylesterase. The consequence is that such contaminated malathion becomes highly toxic to humans because detoxication is inhibited and oxidation becomes important. This led to the poisoning of 2800 workers in Pakistan and the death of 5 (see chap. 5 for metabolism and chap. 7 for more details). 

Tissue esterases have been divided into two classes the A-type esterases, which are insensitive, and the B-type esterases, which are sensitive to inhibition by organo-phosphorus esters. The A esterases include the arylesterases, whereas the B esterases include cholinesterases of plasma, acetylcholinesterases of erythrocytes and nervous tissue, carboxylesterases, lipases, and so on. The nonspecific arylesterases that hydrolyze short-chain aromatic esters are activated by Ca2+ ions and are responsible for the hydrolysis of certain organophosphate triesters such as paraoxon (Figure 10.10B). 

Exposure to some organophosphate cholinesterase inhibitors results in a delayed neuropathy characterized by degeneration of axons and myelin. This effect is not associated with the inhibition of acetylcholinesterase, but rather with the inhibition of an enzyme described as neuropathy target esterase (NTE) however, the exact mechanism of toxicity is not yet fully understood (Munro et al., 1994). For some organophosphate compounds, delayed neuropathy can be induced in experimental animals at relatively low exposure levels, whereas for others the effect is only seen following exposure to supralethal doses when the animal is protected from the acute toxic effects caused by cholinesterase inhibition. 

OP-inhibited acetylcholinesterase and butyrylcholin-esterase are the established biomarkers of OP exposure. The special features that make them good biomarkers are ... 

Tri-ort/io-cresyl phosphate, the contaminant in a homemade liquor called Ginger Jake , which is responsible for delayed neuropathy and paralysis of the legs, is bioactivated to a form that inhibits neuropathy target esterase but not acetylcholinesterase (Casida and Quistad, 2004 Glynn, 2006). Large structures with a 12-20 carbon alkyl chain on the phosphorus atom inhibit fatty acid amide hydrolase but not acetylcholinesterase (Casida and Quistad, 2004). These examples clearly show that OPs which do not affect... 

FIGURE 57.9. Inhibition and aging of serine esterases by diisopropylphosphorofluoridate (DFP). The active site serine is organophosphorylated in the inhibition step. Aging results in net loss of an isopropyl group to yield the monoisopropylphosphoryl esterase. This mode of inhibition and aging has been established for acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and neuropathy target esterase catalytic domain (NEST) (Kropp and Richardson, 2007). 

Milatovic, D., Johnson, M.K. (1993). Reactivation of phosphor-amidated acetylcholinesterase and neuropathy target esterase hy treatment of inhibited enzyme with potassium fluoride. Chem. Biol. Interact. 87 425-30. 

Cohen, J.A., Oosterbaan, R.A., Warringa, M.G.P.J. (1955). The turnover number of ali-esterase, pseudo- and tme cholinesterase and the combination of these enzymes with diisopro-pylfluorophosphonate. Biochim. Biophys. Acta 18 228-35. Coult, D.B., Marsh, D.J., Read, G. (1966). Dealkylation studies on inhibited acetylcholinesterase. Biochem. J. 98 869-73. 

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