Harnessing the catalytic plasticity of the ent-kaurene synthase from Bradyrhizobium japonicum to produce the ent-rosane and ent-pimarane scaffolds

The ent-kaurene synthase catalyses the formation of ent-kaurene through a multistep cyclization process, which is an essential step in the biosynthesis of many bioactive natural products. Channeling the key carbocation intermediates through different reaction pathways offers access to different diterpene scaffolds. Herein, we report that the F72Y mutation of the ent-kaurene synthase from Bradyrhizobium japonicum leads to the formation of tricyclic ent-rosa-5(10),15-diene and ent-pimara-8,15-diene as well as tetracyclic ent-kaurene. The combined computational and experimental studies suggest that Tyr72 serves as a general base for reshaping the catalytic function. This finding provides a promiscuous synthase for further engineering to improve its activity and selectivity, which can be used in the heterologous synthesis of ent-rosane and ent-pimarane diterpenoids.