Mechanistic Insights into Interactions between Bacterial Class I P450 Enzymes and Redox Partners

Abstract

Cytochrome P450 enzymes are highly diversified biocatalysts associated with steroid biosynthesis, xenobiotic metabolism, biosynthesis of natural products, and industrial oxidation reactions. A typical P450 catalytic cycle requires sequential transfer of two electrons from NAD(P)H to the heme-iron reactive center for O2 activation. For the most abundant bacterial Class I P450 systems, this important process is usually mediated by two redox partner proteins including an FAD-containing ferredoxin reductase (FdR) and a small iron–sulfur protein, ferredoxin (Fdx). However, it is often unclear which pair of Fdx and FdR among multiple redox partners is the optimal one for a specific Class I P450 enzyme. To address this important but underexplored question, herein, a reaction matrix network with 16 Fdxs, 8 FdRs, and 6 P450s (against 7 substrates) was constituted. By analyzing the reactivity profiles of 896 P450 reactions, together with phylogenetic analysis, redox potential measurements, structural simulations, and Fdx-P450 molecular docking, we provide important mechanistic insights into the recognition and interactions between bacterial Class I P450 enzymes and redox partners.