Discovery of a Fungal P450 with an Unusual Two-Step Mechanism for Constructing a Bicyclo[3.2.2]nonane Skeleton

Abstract

The successful biomimetic or chemoenzymatic synthesis of target natural products (NPs) and their derivatives relies on enzyme discovery. Herein, we discover a fungal P450 BTG5 that can catalyze the formation of a bicyclo[3.2.2]nonane structure through an unusual twostep mechanism of dimerization and cyclization in the biosynthesis of beticolin 1, whose bicyclo[3.2.2]nonane skeleton connects an anthraquinone moiety and a xanthone moiety. Further investigation reveals that BTG5-T318 not only determines the substrate selectivity but also alters the catalytic reactions, which allows the separation of the reaction to two individual steps, thereby understanding its catalytic mechanism. It reveals that the first heterodimerization undergoes the common oxidation process for P450s, while the second uncommon formal redox-neutral cyclization step is proved as a redox-mediated reaction, which has never been reported. Therefore, this work advances our understanding of P450-catalyzed reactions and paves the way for expansion of the diversity of this class of NPs through synthetic biology.

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“…Biosynthetic studies have identified the fungal cytochrome P450 enzyme, BTG5, responsible for bicyclo[3.2.2]nonane skeleton formation during the biosynthesis of beticolin 1 21 from Cercospora species JNU001. 21 Mechanistic studies revealed a two-step process involving a common P450-catalysed heterodimerisation, followed by an uncommon redox-neutral cyclisation step. The total synthesis of ambruticins, polyketide-derived natural products from the myxobacterium Sorangium cellulosum , has provided standards for investigation of dihydropyran formation during their biosynthesis.…”