Molecular dynamics simulations and free energy calculations have been used to investigate the effect of ligand binding on the enantioselectivity of an epoxide hydrolase (EH) from Aspergillus niger. Despite sharing a common mechanism, a wide range of alternative mechanisms have been proposed to explain the origin of enantiomeric selectivity in EHs. By comparing the interactions of (R)- and (S)-glycidyl phenyl ether (GPE) with both the wild type (WT, E = 3) and a mutant showing enhanced enantioselectivity to GPE (LW202, E = 193), we have examined whether enantioselectivity is due to differences in the binding pose, the affinity for the (R)- or (S)- enantiomers, or a kinetic effect. The two enantiomers were easily accommodated within the binding pockets of the WT enzyme and LW202. Free energy calculations suggested that neither enzyme had a preference for a given enantiomer. The two substrates sampled a wide variety of conformations in the simulations with the sterically hindered and unhindered carbon atoms of the GPE epoxide ring both coming in close proximity to the nucleophilic aspartic acid residue. This suggests that alternative pathways could lead to the formation of a (S)- and (R)-diol product. Together, the calculations suggest that the enantioselectivity is due to kinetic rather than thermodynamic effects and that the assumption that one substrate results in one product when interpreting the available experimental data and deriving E-values may be inappropriate in the case of EHs.

中文翻译：

---

结合对环氧水解酶对映选择性的影响

分子动力学模拟和自由能计算已用于研究配体结合对黑曲霉环氧水解酶（EH）对映选择性的影响。尽管有一个共同的机制，但已经提出了各种各样的替代机制来解释EHs中对映异构体选择性的起源。通过比较（R）-和（S）-缩水甘油基苯基醚（GPE）与野生型（WT，E = 3）和对GPE表现出增强对映选择性的突变体（LW202，E = 193）的相互作用，我们有检查对映选择性是否是由于结合姿势，（R）-或（S）-对映异构体或动力学效应。两种对映体很容易容纳在WT酶和LW202的结合口袋中。自由能的计算表明，两种酶都不对给定的对映异构体优先。在模拟中，两种底物采样了各种各样的构型，其中GPE环氧环的位阻和无阻碳原子都非常接近亲核天冬氨酸残基。这表明替代途径可能导致（S）-和（R）-二醇产品。总之，这些计算表明对映选择性是由于动力学而不是热力学效应引起的，并且在EH的情况下，当解释可用的实验数据并推导E值时，一种底物产生一种产物的假设可能是不合适的。