Rate-limiting step in the decarbamoylation of acetylcholinesterases with large carbamoyl groups.

Carbamates are esters of substituted carbamic acids that react with acetylcholinesterase (AChE) by initially transferring the carbamoyl group to a serine residue in the enzyme active site accompanied by loss of the carbamate leaving group followed by hydrolysis of the carbamoyl enzyme. This hydrolysis, or decarbamoylation, is relatively slow, and half-lives of carbamoylated AChEs range from 4 min to more than 30 days. Therefore, carbamates are effective AChE inhibitors that have been developed as insecticides and as therapeutic agents. In this report, we review recent data showing that decarbamoylation rate constants are independent of the ester leaving group for a series of carbamic acid esters with the same carbamoyl group and that decarbamoylation rate constants decreased by 800-fold when the alkyl substituents on the carbamoyl group increased in size from N-monomethyl- to N,N-diethyl-. We also review data showing that solvent deuterium oxide isotope effects for decarbamoylation decreased from 2.8 for N-monomethylcarbamoyl AChE to 1.1 for N,N-diethylcarbamoyl AChE, indicating a shift in the rate-limiting step from general acid-base catalysis to a likely conformational change in the distorted active site in N,N-diethylcarbamoyl AChE. The nature of such a conformational change is suggested from X-ray crystal structures of AChE phosphorylated by paraoxon.