期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2022
卷号：119
期号：1
DOI：10.1073/pnas.2113770119
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Significance
The completion of the tetracyclic core of etoposide, classified by the World Health Organization as an essential medicine, by the Fe/2OG oxygenase deoxypodophyllotoxin synthase follows a hybrid radical-polar pathway not previously seen in other members of this enzyme class. The implication of a substrate-based benzylic carbocation in this mechanism will inform ongoing efforts to create analogs of this important drug with improved or emergent properties and represents a new route for resolution of the initial substrate radical that is common to members of the class. This study adds to our understanding on a growing number of biochemical transformations in which carbocation intermediates are likely to be crucial.
Deoxypodophyllotoxin contains a core of four fused rings (A to D) with three consecutive chiral centers, the last being created by the attachment of a peripheral trimethoxyphenyl ring (E) to ring C. Previous studies have suggested that the iron(II)- and 2-oxoglutarate–dependent (Fe/2OG) oxygenase, deoxypodophyllotoxin synthase (DPS), catalyzes the oxidative coupling of ring B and ring E to form ring C and complete the tetracyclic core. Despite recent efforts to deploy DPS in the preparation of deoxypodophyllotoxin analogs, the mechanism underlying the regio- and stereoselectivity of this cyclization event has not been elucidated. Herein, we report 1) two structures of DPS in complex with 2OG and (±)-yatein, 2) in vitro analysis of enzymatic reactivity with substrate analogs, and 3) model reactions addressing DPS’s catalytic mechanism. The results disfavor a prior proposal of on-pathway benzylic hydroxylation. Rather, the DPS-catalyzed cyclization likely proceeds by hydrogen atom abstraction from C7', oxidation of the benzylic radical to a carbocation, Friedel–Crafts-like ring closure, and rearomatization of ring B by C6 deprotonation. This mechanism adds to the known pathways for transformation of the carbon-centered radical in Fe/2OG enzymes and suggests what types of substrate modification are likely tolerable in DPS-catalyzed production of deoxypodophyllotoxin analogs.
