Structural regulation of Class-I and Class-II cytochrome P450 enzymes

Project Summary and Abstract Cytochrome P450 enzymes (CYPs) are responsible for a multitude of critical biological transformations, including, but not limited to reactions central to vitamin-D and cholesterol metabolism, steroidogenesis, fatty acid metabolism, and early-stage drug metabolism. In work supported by our ESI MIRA, we addressed three significant questions in this field. First, we demonstrated that proper coordination of substrates, located distal to the active site heme, can influence the molecular recognition of the necessary CYP redox partner proteins along the proximal side of the enzyme. Second, we demonstrated that such long range allostery is bidirectional, thus redox protein recognition can also influence CYP-substrate interactions. Lastly, we demonstrated that the route that constitutes distal-to-proximal allostery is conserved across structurally homologous mammalian and bacterial CYPs. Together, these contributions provide the necessary framework for understanding novel modes of structural regulation in this important class of enzymes. In the proposed MIRA, we will use our biochemical and biophysical experimental workflow to expand these findings into three broad Program Areas. These include i) an investigation of the regulatory potential of mitochondrial CYP-Adrenodoxin encounter complexes, ii) an investigation of drug-induced modulation of CYP-reductase interactions, and iii) elucidation of a pH-sensing mechanism in CYPs. This work represents an expansion of the scope of the ESI MIRA, while also advancing the field by exploring novel modes of structural regulation of CYP function.