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Updated: Dec 20, 2025

A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products
Published on: October 4, 2019
FAD-dependent enzyme-catalysed intermolecular [4+2] cycloaddition in natural product biosynthesis
Lei Gao1,2, Cong Su1, Xiaoxia Du2
1State Key Laboratory of Bioactive Substance and Function of Natural Medicines; CAMS Key Laboratory of Enzyme and Biocatalysis of Natural Drugs, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.
Abstract:
The Diels-Alder reaction is one of the most powerful and widely used methods in synthetic chemistry for the stereospecific construction of carbon-carbon bonds. Despite the importance of Diels-Alder reactions in the biosynthesis of numerous secondary metabolites, no naturally occurring stand-alone Diels-Alderase has been demonstrated to catalyse intermolecular Diels-Alder transformations. Here we report a flavin adenine dinucleotide-dependent enzyme, Morus alba Diels-Alderase (MaDA), from Morus cell cultures, that catalyses an intermolecular [4+2] cycloaddition to produce the natural isoprenylated flavonoid chalcomoracin with a high efficiency and enantioselectivity. Density functional theory calculations and preliminary measurements of the kinetic isotope effects establish a concerted but asynchronous pericyclic pathway. Structure-guided mutagenesis and docking studies demonstrate the interactions of MaDA with the diene and dienophile to catalyse the [4+2] cycloaddition. MaDA exhibits a substrate promiscuity towards both dienes and dienophiles, which enables the expedient syntheses of structurally diverse natural products. We also report a biosynthetic intermediate probe (BIP)-based target identification strategy used to discover MaDA.
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