Acetylcholinesterase carbamylation

Mode of Action. All of the insecticidal carbamates are cholinergic, and poisoned insects and mammals exhibit violent convulsions and other neuromuscular disturbances. The insecticides are strong carbamylating inhibitors of acetylcholinesterase and may also have a direct action on the acetylcholine receptors because of their pronounced stmctural resemblance to acetylcholine. The overall mechanism for carbamate interaction with acetylcholinesterase is analogous to the normal three-step hydrolysis of acetylcholine however, is much slower than with the acetylated enzyme. 

The N-methyl carbamate esters cause reversible carbamylation of the acetylcholinesterase enzyme, allowing accumulation of acetylcholine, the neuromediator substance, at parasympathetic neuroeffector junctions (muscarinic effects), at... 

Carbamates effect the reversible carbamylation of acetylcholinesterase, permitting accumulation of acetylcholine at cholinergic neuroeffector junctions (muscarinic effects), at the myoneural junctions of skeletal muscle, and in the autonomic ganglia (nicotinic effects). CNS function is also impaired. However the relatively large dissociation constant of the carbamyl-enzyme complex indicates that it dissociates more readily than does the organophosphate-enzyme complex, mitigating the toxicity of the carbamate pesticides. The reversibility of the carbamyl-enzyme complex affects (limits) the utility of blood enzyme measurements as a diagnostic tool. 

Carbamates also include pesticides such as Sevin, aldicarb, and carbaryl. They are more degradable than organophosphates and have lower dermal toxicides. Their toxicity is also due to inhibition of acetylcholinesterase but they do not penetrate the central nervous system, so most effects are respiratory in nature. An acetylcholinesterase, which has been carbamylated can undergo spontaneous hydrolysis in vivo, which reactivates the enzyme leading to less severe symptoms or a shorter duration of symptoms. Carbaryl has a low toxicity for mammals however, Perimicarb is highly toxic to mammals, but not readily absorbed through the skin. 

The mode of action of the carbamate insecticides is similar to that of the organophosphates. As shown in Figure 7.15, the reaction yields a carbamylated AChE, followed by decarbamylation via hydrolysis. Carbamates also attack the CNS system, and the symptoms of intoxication are similar to those with the organophosphates. However, unlike the organophosphates, decarbamylation of acetylcholinesterase is rapid, typically in minutes, and therefore carbamate insecticides are regarded as reversible acetylcholinesterase inhibitors. 

These compounds inhibit the hydrolysis of the neurotransmitter acetylcholine by the enzyme acetylcholinesterase within the mammalian nervous system (Zwiener and Ginsburg, 1988). This inhibition causes acetylcholine levels to rise, thus causing cholinergic hyperstimulation at muscarinic and nicotinic receptors. There are important differences in the way carbamates and OPs bind to acetylcholinesterase as well as their abililty to affect the CNS. Carbamates are reversible inhibitors of cholinesterase enzymes. Carbamates create a reversible bond to the cholinesterase enzyme through carbamylation which can spontaneously hydrolyze, reversing toxicity. Carbamate poisoning produces toxicity similar to that of OPs however, the toxicity is usually of a shorter duration and less severe in nature (Lifshitz et al, 1994). In contrast, OPs inhibit cholinesterase via an irreversible bond of phosphate radicals... 

Fig. 2 Seeps involved in Che hydrolysis of acetylcholine (ACh) by acetylcholinesterase (AChE) (I), and in the inhibition of AChE by reversible (II)> carbamyl ester (1I1) and organophosphorus (IV) agents. Heavy, light, and dashed arrows represent extremely rapid. Incecmedlace. and extremely slow or insignificant reactions, respectively. Reproduced from Koelle. G.B.. In The acmacologlc Basis of Therapeutics (Goodman. L.S. and Gilman. A., eds.). Sth ed. Macmillan Publ. Co.. 1975, pg. 448.

Carbamyl choline is is a poor substrate for acetylcholinesterase and nonspecific cholinesterases (74). The nitrate ester (18, Table 2.1)... 

S-(-f)-Acetyl-j3-methylcholine (the euto-mer) is hydrolyzed by acetylcholinesterase at about half the rate of acetylcholine the R-(-)-enantiomer is a weak inhibitor of the enzyme (82). Work of Ringdahl (85) suggests that the markedly lower pharmacological activity of R-(-)-acetyl-j3-methylcholine is a result both of lower affinity for the muscarinic receptor(s) and lower intrinsic activity. The antipodes of carbamyl-jS-methylcholine (bethanechol) (34) displayed a eudismie ratio of 740 at rat jeju-... 

Acetylcholinesterase (AChE) inhibitors are successfully used as drugs for the treatment of Alzheimer s disease. Physostigmine, also named eserine, has been isolated from the Calabar bean Physostigmina Venenosum). It is classified as a pseudo-irreversible inhibitor because it reacts with acetyl- as well as butyryl-cholinesterase (BChE) to form a carbamylated serine which is hydrolyzed again with a fi/2 of 40 min [73]. Physostigmine inhibits AChE with an IC50 of 27.9 nM and BChE with an IC50 of 16 nM. Its enantiomer is 350 times less active on AChE and 150 times less active on BchE [74]. 

Acetylcholinesterase. Altered acetylcholinesterase less sensitive to organophosphorus and carbamate insecticides has been observed in a wide variety of insects and mites (51). Acetylcholinesterase inhibiting insecticides phosphorylate or carbamylate the serine residue in the active site of the enzyme preventing vital catalysis of acetylcholine. Resistance due to reduced sensitivity to inhibition of this target enzyme has been found in house fly, mosquitoes, green rice leafhopper, and both phytophagous and predacious species of mites. 

Carbary I (1-naphthyl methylcarbamate) is a chemical in the carbamate family used chiefly as an insecticide. It is a colorless white crystalline solid. Carbaryl disrupts the nervous system by adding a carbamyl moiety to the active site of the acetylcholinesterase enzyme, which prevents it from interacting with acetylcholine.1 It is classified as a likely human carcinogen by the EPA. The pesticide is used indiscriminately, so the toxicity has raised public concern about the ecosystem and human health. Carbaryl is lethal to many non-target insects such as the honeybee. Accumulation of the pesticide occurs in many aquatic organisms such as catfish and algae.2 Due to public health and ecosystem concerns a number of analytical procedures have been used to determine carbaryl concentrations. 

Butyrylcholinesterase occurs in the liver and at the motor endplates in muscle fibers and at synapses together with aeetylcholinesterase (Silver 1974). It is estimated that approximately 15% of total cholinesterase activity in the nervous system is due to the nonspecific cholinesterase activity in some of the white matter (Ecobichon and Joy 1982), and the synthesis of this nonspecific cholinesterase occurs in the liver. Several organophosphorus compounds can react and phosphorylate acetylcholinesterase carbamate ester compounds can carbamylate the enzyme, so both can inhibit the acetylcholinesterase. 

The carbamyl ester inhibitors compete with acetylcholine for the active site of the enzyme. Acetylcholinesterase becomes carbamylated as it cleaves the ester linkage. The carbamyl group prevents acetylcholine from binding to the active site of acetylcholinesterase for a period of minutes to hours. Upon decarbamylation, the enzyme regains its ability to cleave acetylcholine. 

Wilson IB, Harrison MA, Ginsberg S. Carbamyl derivatives of acetylcholinesterase. J Biol Chem 1961 236 1498-1500. 

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