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Butyrylcholinesterase (BuChE) is a stoichiometric bioscavenger against organophosphorus (OP) nerve agent poisoning, and efforts to make BuChE variants that are catalytically active against a wide spectrum of nerve agents have been ongoing for the last decade. In order to understand the structural consequences for BuChE, we carried out extensive molecular dynamics (MD) simulations on wild-type BuChE (PDB ID: 1P0I) and several known and new variants of this enzyme, but without the presence of any ligand in the active site. The MD simulations on WT-BuChE identified two labile orientations for the catalytic serine, and also showed the likelihood of a backdoor. Upon changes at the G116 position, severe alterations around the active site region were identified. Simulations on both G117H and G117N variants showed the existence of a bound water molecule that is in close proximity to S198. Modeling of the E197Q mutant suggested that Q197 can be in two distinct orientations, one similar to the E202Q-AChE crystal structure and another in proximity to G439 and E441. The double mutant, G117H/E197Q, was found to have structural characteristics of both G117H and E197Q. In light of the computational results, previous experimental observations are discussed., (Copyright (c) 2010 Elsevier Ireland Ltd. All rights reserved.)

The phosphonylation mechanism of AChE and the S203C mutation by sarin (GB) is evaluated using two reaction schemes: a small model nucleophile (ethoxide, CH(3)CH(2)O(-)) and quantum mechanical/molecular mechanical (QM/MM) simulations. Calculations utilizing small model nucleophiles indicate that the reaction barrier for addition to GB is the rate-limiting step for both ethoxide and ethyl thiolate (CH(3)CH(2)S(-)); moreover, the activation barrier for addition to the phosphorus center of GB by ethyl thiolate is significantly larger (13.2 kcal/mol) than for ethoxide (8.3 kcal/mol). The decomposition transition state for both nucleophiles was determined to be approximately 1 kcal/mol. QM/MM simulations for AChE suggest a similar reaction mechanism for phosphonylation of the catalytic S203; however, the relative energetics are altered significantly compared to the isolated system. QM/MM results indicate that formation of the penta-coordinate intermediate is the rate-limiting step in the enzymatic system, with an activation barrier of 3.6 kcal/mol. Hydrogen-bonding interactions between the fluoride leaving group of GB with Y124 in AChE are observed throughout the reaction profile. The S203C mutation alters the relative energetics of the reaction, increasing the energy barrier for formation of the penta-coordinate intermediate to a value of 4.5 kcal/mol; moreover, the penta-coordinate intermediate (as product) is stabilized by an additional 6 kcal/mol when compared to wild-type AChE., (Copyright (c) 2010 Elsevier Ireland Ltd. All rights reserved.)

Density functional theory calculations were employed to study the reaction of five nerve agents with model nucleophiles, including EtX(-) and EtXH (X=O, S, Se) for serine, cysteine and selenocysteine, respectively. Calculations at the B3LYP/6-311++G(2d,p) level of theory predict an exothermic reaction between ethoxide and all of the nerve agents studied. As compared to EtO(-) as a nucleophile, these reactions become approximately 30 kcal/mol more endothermic for EtS(-), and by approximately 40 kcal/mol for EtSe(-). The equivalent reactions with the neutral nucleophiles (EtXH) were more endothermic. The effect of solvation on the reaction thermochemistry was determined using a polarizable continuum model simulating the dielectric constant of chloroform. While there was a large exothermic shift for reactions involving charged nucleophiles with solvation modeling, the corresponding shift was minimal for the reaction with neutral nucleophiles.