Transition States in a Protein Environment − ONIOM QM:MM Modeling of Isopenicillin N Synthesis

To highlight the role of the protein in metal enzyme catalysis, we optimize ONIOM QM:MM transition states and intermediates for the full reaction of the nonheme iron enzyme isopenicillin N synthase (IPNS). Optimizations of transition states in large protein systems are possible using our new geometry optimizer with quadratic coupling between the QM and MM regions [Vreven, T. et al. <i>Mol. Phys.</i> <b>2006</b>, <i>104</i>, 701−704]. To highlight the effect of the metal center, results from the protein model are compared to results from an active site model containing only the metal center and coordinating residues [Lundberg, M. et al. <i>Biochemistry</i> <b>2008</b>, <i>47</i>, 1031−1042]. The analysis suggests that the main catalytic effect comes from the metal center, while the protein controls the reactivity to achieve high product specificity. As an example, hydrophobic residues align the valine substrate radical in a favorable conformation for thiazolidine ring closure and contribute to product selectivity and high stereospecificity. A low-barrier pathway for β-lactam formation is found where the proton required for heterolytic O−O bond cleavage comes directly from the valine N−H group of the substrate. The alternative mechanism, where the proton in O−O bond cleavage initially comes from an iron water ligand, can be disfavored by the electrostatic interactions with the surrounding protein. Explicit protein effects on transition states are typically 1−6 kcal/mol in the present enzyme and can be understood by considering whether the transition state involves large movements of the substrate as well as whether it involves electron transfer.