Thermodynamic, Electronic, Structural, and Kinetic Characterizations of Cytochrome P450 Compounds I & II

University Of California-Irvine, Irvine CA

Investigators

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Abstract

Project Summary The research outlined in this proposal seeks to further our understanding of the factors that govern C-H bond activation in cytochrome P450 catalysis. Over the past several years my group has made significant contributions to this area. Our results have impacted not only the way people think about P450 catalysis but also metal-oxo mediated C-H bond activation in general. We have led the way in the capture and characterization of critical intermediates in the P450 catalytic cycle and developed theories to describe how Nature biases enzymes for C-H bond activation. Still, much remains to be done. Our understanding of the factors that govern C-H bond activation in P450s remains incomplete. Importantly, results from our last funding period have shown that P450 can serve as a platform from which to attack some of the most important and fundamental questions in the field of C-H bond activation. There remains a debate in the field about the factors that govern reactivity in metal-oxo driven C-H bond activation. The debate centers on whether ground state thermodynamics, in the form of bond strengths, play the dominant role in determining reactivity or whether some other property of the system (e.g., spin density on the oxo ligand, metal-oxo basicity, access to low-lying excited states, or enhanced H-atom tunneling facilitated by strong electron donation from the axial ligand) can provide an intrinsic lowering of the activation barrier. The examination of this fundamental issue has been hindered not only by the difficulty of measuring these quantities for reactive high-valent species but also by the lack of a series of isoelectronic and isostructural compounds over which these quantities can be varied. We have shown that P450 can fill this void. During the previous funding period we expanded the range of P450s in which we can prepare compounds I and II for experimental studies. We now have access to an isoelectronic series of P450 intermediates with varying axial ligation. This and other innovations will allow us to measure the ground state thermodynamics of C-H bond cleavage, quantify the degree of H-atom tunneling and ferryl basicity, determine the amount of oxyl-radical character in compound I, examine the accessibility of low-lying excited states, and, importantly, track how these quantities (and the reactivity towards C-H bonds) change as a function of electron donation from the axial ligand. There is currently no other system, synthetic or biological, that allows for a similar set of measurements and discovery.