We present the first detailed characterization of the electronic structure of P450 compound I (P450-I). P450s are thiolate-ligated heme proteins that excel at the activation of inert C-H bonds. In humans, these enzymes are responsible for the phase I metabolism of ~75% of all pharmaceuticals. A major goal of bioinorganic chemistry has been to elucidate factors that enable cytochrome P450 to activate inert hydrocarbons1. Central to these efforts have been attempts to define the geometric and electronic structures of the highly reactive intermediate, termed compound I, in which the ferryl iron (S=1) is antiferromagnetically coupled to a radical (S=1/2). Recently, our laboratory made a breakthrough on this front, discovering how to prepare P450-I in high yield2. This discovery has opened the door for investigations that could provide key insights into the factors that govern C-H bond activation.
We have characterized the electronic structure of P450 compound I utilizing advanced pulse EPR methods. We have resolved 33S(Cys), 1H(Cys) and 14N (porphyrin) hyperfine coupling interactions, which are the first-hand reporters for the degree of delocalization of the radical between the porphyrin and the axial thiolate ligand. The measurement of electronic relaxation rates has allowed us to determine the magnitude of the exchange interaction between ferryl and the ligand radical as well as the ferryl zero-field splitting parameters. Both of these parameters have key implications for the validity of the two-state reactivity mechanism, proposed earlier.3
The resolved properties are compared with those obtained for a structurally similar moiety of chloroperoxidase (CPO), which, however, cannot oxidize unactivated hydrocarbons. This comparison provides an important basis for understanding the factors influencing the reactivity of the thiolate-ligated heme iron sites.
1. I.G.Denisov, et al.//Chem.Rev. 105:2253-2277(2005)
2. J.Rittle, M.T.Green//Science 330(6006):933-937(2010)
3. H.Hirao, et al.//J.Am.Chem.Soc 128:8590-8606(2006)