Mechanistic Insights into the Decoupled Desaturation and Epoxidation Catalyzed by Dioxygenase AsqJ Involved in the Biosynthesis of Quinolone Alkaloids

Abstract

AsqJ from Aspergillus nidulans is a nonheme FeII/α-ketoglutarate-dependent dioxygenase that catalyzes the conversion of benzodiazepinedione into 4′-methoxyviridicatin, which is a key step in the biosynthesis of quinolone alkaloids. A series of recent experiments have demonstrated that AsqJ is able to perform the decoupled desaturation and epoxidation reactions. Herein, on the basis of the published crystal structures, combined quantum mechanics and molecular mechanics (QM/MM) calculations have been performed to explore both the desaturation and epoxidation processes. Our calculations reveal that the quintet state of the FeIV–O complex is the ground state, and the catalytic reaction occurs on the quintet-state surface. The FeIV–oxo species should first undergo an isomerization to initiate the reactions. In the desaturation process, the abstraction of the first hydrogen atom is suggested to follow the σ-channel mechanism. This step is calculated to be rate-limiting with an energy barrier of 19.3 kcal/mol. The abstraction of the second hydrogen atom is found to be quite easy. After the desaturation process, the regenerated FeIV–oxo species first attacks the C═C bond of the desaturated intermediate to form a carbon-based radical intermediate, corresponding to an energy barrier of 18.1 kcal/mol, then the radical intermediate completes the ring closure with a barrier of 3.9 kcal/mol. Besides, the calculations using the substrate analogous that lacks the N4-methyl reveal that the H atom abstraction by FeIV–oxo is still accessible, which suggests that the absence of N4-methyl does not affect the desaturation process itself but may influence the other processes that occur prior to the desaturation.