Computational Investigation of Cytochrome P450-Catalyzed Oxetane Formation in Taxol Biosynthesis

Paclitaxel (Taxol) is widely recognized as one of the most effective natural anticancer drugs, yet its production is limited by its scarce natural supply and the low yields obtained from its chemical synthesis. One of the most puzzling steps in its biosynthesis is the formation of the oxetane-containing D-ring, a key structural element necessary for its microtubule binding and anticancer activity. In our study, we use density functional theory (DFT) calculations to explore how cytochrome P450 drives this critical transformation. We focus on two proposed reaction pathways─the ene-acetoxy (EA) pathway and the ene-hydroxy (EH) pathway─and investigate how they operate on both the high-spin (quartet) and low-spin (doublet) states of Compound I. Our results indicate that on the high-spin surface, epoxidation occurs in two steps, with the first step being the slow, rate-limiting part of the reaction, while on the low-spin surface, the process proceeds in a single step. The computed activation energies and the observed stability of the epoxide intermediates support the idea that enzyme-catalyzed epoxidation is a pivotal intermediate in the formation of the oxetane ring during Taxol biosynthesis. These mechanistic insights not only enhance our understanding of this complex biosynthetic pathway but also have the potential to guide future efforts aimed at improving Taxol production.