Application of a SCC-DFTB QM/MM approach to the investigation of the catalytic mechanism of fatty acid amide hydrolase

Abstract

Self-consistent charge density functional tight binding (SCC-DFTB) is a promising method for hybrid quantum mechanics/molecular mechanics (QM/MM) simulations of enzyme-catalyzed reactions. The acylation reaction of fatty acid amide hydrolase (FAAH), a promising drug target, was investigated by applying a SCC-DFTB/CHARMM27 scheme. Calculated potential energy barriers resulted in reasonable agreement with experiments for oleamide (OA) and oleoylmethyl ester (OME) substrates, outperforming previous calculations performed at the PM3/CHARMM22 level. Furthermore, the experimental preference of FAAH in hydrolyzing OA faster than OME was adequately reproduced by calculations. All these findings indicate that the SCC-DFTB/CHARMM27 approach can be successfully applied to mechanistic investigations of FAAH-catalyzed reactions.

Anandamide Hydrolysis in FAAH Reveals a Dual Strategy for Efficient Enzyme-Assisted Amide Bond Cleavage via Nitrogen Inversion

“…This process has been shown to be slightly favored (∼2 kcal mol −1 ) with respect to the subsequent formation of the acyl−enzyme (B C), suggesting a concerted acylation step of oleamide hydrolysis in FAAH. 19,23,25 These results agree also with the study of Tubert-Brohman et al 30 who, through Monte Carlo QM/MM calculations at the semiempirical PDDG/PM3 level, confirmed a concerted mechanism for the acylation process for the oleoyl substrates. Lodola et al also investigated the deacylation of FAAH, acylated by oleamide and few carbamate and urea-based covalent inhibitors.…”

“…57 Therefore, given the lower calculated free energy barrier for the deacylation reaction (∼16 kcal mol −1 ), with respect to the acylation reaction (∼19 kcal mol −1 ), the latter can be considered to be the rate-determining step of the entire chemical process, in agreement with previous studies. 19,[21][22][23][25][26][27]30 Overall, our results indicate a single-step mechanism for both acylation and deacylation reactions in FAAH. This is a highly concerted mechanism, where spontaneous and complete proton transfers without stepwise intermediate states facilitate the nucleophilic attacks and leaving group departures.…”

A Catalytic Mechanism for Cysteine N-Terminal Nucleophile Hydrolases, as Revealed by Free Energy Simulations

“…Although SCC-DFTB method has some limitations, it has become increasingly widely applied to biological problems [40], and it has been reported to provide results in agreement with DFT calculations [43], [44]. Furthermore, its efficiency made it feasible to sample millions of conformations for enzyme systems and to determine the free energy profiles of the enzyme-catalyzed reactions [45], complementing results obtained with ab initio free energy methods [46], [47].…”

Computational insights into function and inhibition of fatty acid amide hydrolase

“…Regarding the acylation step, Mulholland et al emphasized that one key aspect of the FAAH catalytic machinery is the pyramidal inversion of the substrate nitrogen, which is occurring during the enzyme acylation [114,115]. Indeed, this event properly orients the nitrogen lone pair toward Ser217 for the protonation and subsequent exit of the leaving group (Scheme 2).…”

“…the irreversible inhibitor methyl arachidonyl fluorophosphonate (MAFP), prompted several computational investigations on both FAAH catalysis and inhibition. Much credit for important computational insights is due to the groups of Jorgensen [28,32,113] and Mulholland [29e31,39,40,114,115], who have independently modeled the acylation step of the enzymatic catalysis (A / D in Scheme 2) using different computational approaches.…”