Combination of Molecular Modeling strategies to reveal why Carboxylesterases subtypes 1 and 2 metabolized different substrates

Seminario; Programa Pós-graduação de Ciências Farmacêuticas da UFRGS; 2022

Universidade Federal Do Rio Grande Do Sul

Human carboxylesterases (CES) play an important role in the metabolization/biotransformationof several endogenous and exogenous substrates, with subtypes CES1 and CES2 as the most important members, such is the case for the esterification levels of endogenous lipids and the metabolization and bioactivation of numerous approved drugs and prodrugs. During the last years, several efforts have been made to elucidate the different substrate specificity between the CES1 and CES2, but several inconsistencies are still without explanation. In this study, the development of a methodology, using a combination of classical molecular mechanics (MM) and hybrid molecular mechanic/quantum mechanics (QM/MM) simulation strategies, were performed in order to unveiled the differences between CES1 and CES2 that produced the distinct hydrolysis substrate pattern. For developing this methodology, a set of already reported CES1 and CES2 substrates, $p$-nitrophenyl esters (NPE), were used.The initial molecular structure comparison analysis showed that CES2 displayed a 80~\AA\textsuperscript{3} higher volume area on its active site cavity than the respective CES1, because of the presence of a loop on the entrance of CES1 active site cavity. The free-energy binding interaction analysis exhibited a correlation between the free-energy van del Waals (VDW) component of interaction and the reported experimental ($K\textsubscript{m}$) for the 5 studied NPE with both CES, originated from the hydrophobic interaction between the different alkyl NPE moieties and the residues present on tunnel section of the active site cavity. The hybrid QM/MM simulation performed on the hydrolysis reaction of two NPE (pNPP and pNPTA) by CES1 and CES2 showed that the rate-limiting step in the studied reactions was the second transition state (TS\textsubscript{2}), corresponding to the deacylation process. A relationship was observed between the free-energies of reaction involved in the hydrolysis reaction of pNPP and pNPTA and the corresponding reported experimental $k\textsubscript{cat}$ for CES1 and CES2, originated on the steric hindrance of the NPE TS\textsubscript{2} structure during the course of the hydrolysis reaction. The developed methodology showed high efficiency to reveal the molecular features behind the catalytic differences between CES1 and CES2 to produced the hydrolysis of a series of already reported NPE substrates. This developed methodology would be used on future analysis of reported and approved drugs and prodrugs.