Computer simulations of acetylcholinesterase inhibition by nerve agent soman

Abstract

Acetylcholinesterase (AChE) is a crucial enzyme in the cholinergic nervous system that hydrolyses neurotransmitter acetylcholine (ACh) and terminates synaptic signals by reducing the effective concentration of ACh in the synaptic clefts. The catalytic serine of AChE can be phosphonylated by organophosphate compounds such as snake venoms, insecticides and nerve agents (nerve toxins) and form stable covalent enzyme-inhibitor adducts. These covalent adducts can either be reactivated by nucleophilic compounds with oxime functional units to some limited extent or undergo a very fast dealkylation reaction, also termed "aging". This phosphonylation and aging process leads to irreversible AChE inhibition, results in accumulation of excess ACh at the synaptic clefts and causes neuromuscular paralysis. By employing Born-Oppenheimer ab initio QM/MM molecular dynamics simulations with umbrella sampling, a state-of-the-art approach to simulate enzyme reactions, we have characterized the phosphonylation reaction mechanism between AChE and soman as well as the aging mechanism of soman phosphonylated AChE and determined their free energy profiles. The phosphonylation reaction between AChE and soman is an associative nucleophilic substitution mechanism that is initiated when the nucleophilic Ser200 attacks soman's phosphorus atom, with His440 acting as a general base. In the elimination step, Try121 of the catalytic gorge forms hydrogen bonds with the leaving fluorine atom prior to its dissociation from the active site. Once a stable covalent adduct forms, the aging reaction begins with scission of the alkoxyl covalent bond, connecting the bound soman's alkyl group to the phosphonate moiety. Cleavage of the alkyl chain is swiftly followed with a methyl group rearrangement of the alkyl group resulting in a stable tertiary carbenium, which is hydrated to tertiary alcohol by a reactive water facilitated by Tyr121. Our characterized mechanisms and simulation results provide new detailed insights into this biologically important process. Mechanistic details are of significant interest for the development of novel strategies to reduce the toxic effects of soman by facilitating the search and design of novel compounds capable of slowing the aging of nerve agent inhibited AChEs as well as the design of effective reactivators for the aged conjugates. Most importantly, this work would facilitate the design of catalytic enzymes capable of hydrolyzing nerve agents