Cytochromes P450 (P450s) are heme-containing enzymes known for catalyzing hydroxylation reactions with exceptional regioselectivity. Central to their mechanism is Compound I, a fleeting transition state during turnover, which exists as an iron(IV) oxo complex with an additional oxidative equivalence delocalized over the heme ligand scaffold and axial cysteine ligand. In an effort to understand how protein scaffolds stabilize and direct the activity of these oxidative species, we computationally designed a de novo Fe-corrole binding protein. Corroles are one carbon contracted porphyrinoids that are known to help stabilize high valent metals and can hold an oxidative equivalent making them an excellent starting point to access oxidizing species related to P450 turnover.
The process began with constructing a simple four alpha-helix coiled-coil backbone that was iteratively refined into a single chain protein capable of housing a bulky metallocofactor (FeTPC(Cl), TPC=5,10,15-triphenylcorrole) coordinated by an axial histidine.. The protein was successfully expressed in E.Coli and shown to bind to the corrole by UV-vis spectroscopy. Initial data has demonstrated guaiacol oxidation with hydrogen peroxide. Using diffusion-based design tools we are expanding on this designed backbone to incorporate a substrate binding site to better direct the reactivity of the anchored metal center.