Nitrile Hydratases and Epoxide-Transforming Enzymes: Quantum Chemical Modeling of Reaction Mechanisms and Selectivities

Quantum chemical studies of enzymatic reactions are able to provide detailed insight into mechanisms and catalytic strategies. The energetic feasibility of proposed mechanisms can be established, and new possible reaction pathways can be put forward. The role of the involved active site residues can be analyzed in detail and the origins for experimentally observed selectivities can be investigated. Density functional theory (DFT), in particular the hybrid functional B3LYP, is the method of choice in this kind of studies. In this thesis, the reaction mechanisms of several enzymes have been explored using the B3LYP functional. The studied enzymes include limonene epoxide hydrolase (LEH), soluble epoxide hydrolase (sEH), haloalcohol dehalogenase (HheC), and nitrile hydratase (NHase). Transition states and intermediates along various reaction pathways were optimized and evaluated. For the three epoxide-transforming enzymes, the role of the proposed catalytic residues could be confirmed. Analysis of in silico mutations helped to quantify the effect of various functional groups on the barriers and regioselectivities of epoxide opening. A detailed analysis of the factors governing the enzymatic regioselectivities is given. For nitrile hydratase, various putative firstand second-shell mechanisms have been studied. Active site models based on both the Co(III)-NHase and the Fe(III)-NHase were employed. The studied mechanisms include general base-catalyzed reaction pathways with water as nucleophile as well as two pathways involving cysteinesulfenate as nucleophile. Several computed mechanisms exhibit similar barriers, making it difficult to pinpoint the true NHase mechanism.