Density Functional Theory / Nudged Elastic Band Investigation of the Hydroxylation Reaction Mechanism Catalyzed by P450cam

We have calculated the complete minimum-energy reaction path for the hydroxylation of camphor by the P450 enzyme from Pseudomonas putida using the nudged elastic band method of Jonsson and co-workers[1]. Single-point force and energy calculations on pathway images were performed at the hybrid density functional level of theory (B3LYP) with large basis sets for the iron atom (6-311+G) and O$_{2}$ ligand (6-31+G*) on a ~100 atom active site extracted from a recent high-resolution crystal structure[2]. Our model includes the heme group liganded to both Cys357 and dioxygen and we also include Thr251 and Asp252, which have been shown to significantly affect product yield by mutational studies[3]. We find that, upon transfer of the 2nd electron to the active site, the Fe-O$_{2}$ moiety is unstable and decays to a Fe-OOH- intermediate via a Asp252-H$_{2}$O proton transfer chain. The barrier for dioxygen cleavage and the identity of the reactive species will be discussed. [1] H. Jonsson, G. Mills, K.W. Jacobsen, in Classical and Quantum Dynamics in Condensed Phase Simulation, World Scientific (1998). [2] I. Schlichting, et al., Science 287, no. 5458, p. 1615-1622 (2000). [3] R. Davydov, et al., J. Am. Chem. Soc., 123: 1403-1415 (2001).