Catalytic Reaction Mechanism of Acetylcholinesterase Determined by Born−Oppenheimer Ab Initio QM/MM Molecular Dynamics Simulations

Abstract

Acetylcholinesterase (AChE) is a remarkably efficient serine hydrolase responsible for the termination of impulse signaling at cholinergic synapses. By employing Born-Oppenheimer molecular dynamics simulations with B3LYP/6-31G(d) QM/MM potential and the umbrella sampling method, we have characterized its complete catalytic reaction mechanism for hydrolyzing neurotransmitter acetylcholine (ACh) and determined its multi-step free energy reaction profiles for the first time. In both acylation and deacylation reaction stages, the first step involves the nucleophilic attack to the carbonyl carbon with the triad His447 serving as the general base, and leads to a tetrahedral covalent intermediate stabilized by the oxyanion hole. From the intermediate to the product, the orientation of His447 ring needs to be adjusted very slightly, and then the proton transfers from His447 to the product and the break of the scissile bond happen spontaneously. For the three-pronged oxyanion hole, it only makes two hydrogen bonds with the carbonyl oxygen at either the initial reactant or the final product state, but the third hydrogen bond is formed and stable at all transition and intermediate states during the catalytic process. At the intermediate state of the acylation reaction, a short and low-barrier hydrogen bond (LBHB) is found to be formed between two catalytic triad residues His447 and Glu334, and the spontaneous proton transfer between two residues has been observed. However, it is only about 1 ~ 2 kcal/mol stronger than the normal hydrogen bond. In comparison with previous theoretical investigations of the AChE catalytic mechanism, our current study clearly demonstrates the power and advantages of employing Born-Oppenheimer ab initio QM/MM MD simulations in characterizing enzyme reaction mechanisms.

Why Does the G117H Mutation Considerably Improve the Activity of Human Butyrylcholinesterase against Sarin? Insights from Quantum Mechanical/Molecular Mechanical Free Energy Calculations

“…In the QM/MM-FE method, the first-principles QM/MM reaction-coordinate calculations were followed by free-energy perturbation (FEP) calculations in order to more reasonably account for the dynamic effects of the protein environment on the free energy barriers for the enzymatic reactions. Our QM/MM simulations are based on the pseudobond first-principles QM/MM approach, (29–32) which has been demonstrated to be a powerful tool in simulating a variety of enzymes, (28, 33–50) and some theoretical predictions (39, 49, 51) were subsequently confirmed by the experimental studies. (35, 49, 52–53) The computational results clearly reveal the reaction mechanisms and free energy profiles of the reactivation reactions of sarin-phosphorylated wild-type human BChE and the G117H mutant.…”

Aging Mechanism of Soman Inhibited Acetylcholinesterase

“…In the present work, we filled this gap and characterized the aging mechanism of soman phosphonylated AChE by employing Born Oppenheimer ab initio QM/MM molecular dynamics simulations with umbrella sampling method, an approach that has been extensively applied to study enzyme reactions 8,50–59 . The characterized aging mechanism is schematically illustrated in Figure 4.…”

Reaction Pathway and Free Energy Profiles for Butyrylcholinesterase-Catalyzed Hydrolysis of Acetylthiocholine

“…40 However, a series of recent studies demonstrated that at a low substrate concentration, hydrolysis of ACh is rate-determined by the acylation reaction, 2, 41, 46 which is consistent with our recently reported study. 47 Despite extensive studies on the catalytic mechanisms of cholinesterase-catalyzed hydrolysis of ACh/ATCh, 1-2, 15, 27-30, 33-34, 36, 38, 48-50 questions remain for the mechanism of the substrate activation for BChE-catalyzed hydrolysis of ACh/ATCh. For example, what is the main factor leading to the observed substrate activation effect?…”