Beyond the cyclopropyl ring formation: fungal Aj_EasH catalyzes asymmetric hydroxylation of ergot alkaloids

Ergot alkaloids (EAs) are among the most important bioactive natural products. Fe II / α -ketoglutarate-dependent dioxygenase Aj_EasH from Aspergillus japonicus is responsible for the formation of the cyclopropyl ring of the ergot alkaloid (EA) cycloclavine ( 4 ). Herein we reconstituted the biosynthesis of 4 in vitro from prechanoclavine ( 1 ) for the first time. Additionally, an unexpected activity of asymmetric hydroxylation at the C-4 position of EA compound festuclavine ( 5 ) for Aj_EasH was revealed. Furthermore, Aj_EasH also catalyzes the hydroxylation of two more EAs 9,10-dihydrolysergol ( 6 ) and elymoclavine ( 7 ). Thus, our results proved that Aj_EasH is a promiscuous and bimodal dioxygenase that catalyzes both the formation of cyclopropyl ring in 4 and the asymmetric hydroxylation of EAs. Molecular docking (MD) revealed the substrate-binding mode as well as the catalytic mechanism of asymmetric hydroxylation, suggesting more EAs could potentially be recognized and hydroxylated by Aj_EasH. Overall, the newly discovered activity empowered Aj_EasH with great potential for producing more diverse and bioactive EA derivatives. Key points • Aj_EasH was revealed to be a promiscuous and bimodal Fe II /α-ketoglutarate-dependent dioxygenase . • Aj_EasH converted festuclavine, 9,10-dihydrolysergol, and elymoclavine to their hydroxylated derivatives . • The catalytic mechanism of Aj_EasH for hydroxylation was analyzed by molecular docking . Graphical abstract