Fungal P450 Deconstructs the 2,5-Diazabicyclo[2.2.2]octane Ring <i>En Route</i> to the Complete Biosynthesis of 21<i>R</i>-Citrinadin A
Shuai Liu, Qiuyue Nie, Zhiwen Liu, Siddhant Patil, Xue Gao
Abstract
Prenylated indole alkaloids (PIAs) possess great structural diversity and show biological activities. Despite significant efforts in investigating the biosynthetic mechanism, the key step in the transformation of 2,5-diazabicyclo[2.2.2]octane-containing PIAs into a distinct class of pentacyclic compounds remains unknown. Here, using a combination of gene deletion, heterologous expression, and biochemical characterization, we show that a unique fungal P450 enzyme CtdY catalyzes the cleavage of the amide bond in the 2,5-diazabicyclo[2.2.2]octane system, followed by a decarboxylation step to form the 6/5/5/6/6 pentacyclic ring in 21 R -citrinadin A. We also demonstrate the function of a subsequent cascade of stereospecific oxygenases to further modify the 6/5/5/6/6 pentacyclic intermediate en route to the complete 21 R -citrinadin A biosynthesis. Our findings reveal a key enzyme CtdY for the pathway divergence in the biosynthesis of PIAs and uncover the complex late-stage post-translational modifications in 21 R -citrinadin A biosynthesis.