Quantum Mechanical/Molecular Mechanical Free Energy Simulations of the Glutathione S-Transferase (M1-1) Reaction with Phenanthrene 9,10-Oxide

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• 作者：Lars Ridder ; Ivonne M. C. M. Rietjens ; Jacques Vervoort ; and Adrian J. Mulholland
• 刊名：Journal of the American Chemical Society
• 出版年：2002
• 出版时间：August 21, 2002
• 年：2002
• 卷：124
• 期：33
• 页码：9926 - 9936
• 全文大小：150K
• 年卷期：v.124,no.33(August 21, 2002)
• ISSN：1520-5126

文摘

Glutathione S-transferases (GSTs) play an important role in the detoxification of xenobiotics inmammals. They catalyze the conjugation of glutathione to a wide range of electrophilic compounds.Phenanthrene 9,10-oxide is a model substrate for GSTs, representing an important group of epoxidesubstrates. In the present study, combined quantum mechanical/molecular mechanical (QM/MM) simulationsof the conjugation of glutathione to phenanthrene 9,10-oxide, catalyzed by the M1-1 isoenzyme from rat,have been carried out to obtain insight into details of the reaction mechanism and the role of solvent presentin the highly solvent accessible active site. Reaction-specific AM1 parameters for sulfur have been developedto obtain an accurate modeling of the reaction, and QM/MM solvent interactions in the model have beencalibrated. Free energy profiles for the formation of two diastereomeric products were obtained frommolecular dynamics simulations of the enzyme, using umbrella sampling and weighted histogram analysistechniques. The barriers (20 kcal/mol) are in good agreement with the overall experimental rate constantand with the formation of equal amounts of the two diastereomeric products, as experimentally observed.Along the reaction pathway, desolvation of the thiolate sulfur of glutathione is observed, in agreement withsolvent isotope experiments, as well as increased solvation of the epoxide oxygen of phenanthrene 9,10-oxide, illustrating an important stabilizing role for active site solvent molecules. Important active siteinteractions have been identified and analyzed. The catalytic effect of Tyr115 through a direct hydrogenbond with the epoxide oxygen of the substrate, which was proposed on the basis of the crystal structureof the (9S,10S) product complex, is supported by the simulations. The indirect interaction through a mediatingwater molecule, observed in the crystal structure of the (9R,10R) product complex, cannot be confirmed toplay a role in the conjugation step. A selection of mutations is modeled. The Asn8Asp mutation, representingone of the differences between the M1-1 and M2-2 isoenzymes, is identified as a possible factor contributingto the difference in the ratio of product formation by these two isoenzymes. The QM/MM reaction pathwaysimulations provide new and detailed insight into the reaction mechanism of this important class of detoxifyingenzymes and illustrate the potential of QM/MM modeling to complement experimental data on enzymereaction mechanisms.