The microbial type and multiproduct sesquiterpene synthase RlMTPSL4 from the liverwort Radula lindenbergiana produces the new sesquiterpene hydrocarbon 4,5-diepi-isoishwarane as the main product, besides the known compounds germacrene A, α-selinene, eremophilene and 4,5-diepi-aristolochene. The enzyme mechanism for the formation of the main product and its absolute configuration were determined through isotopic labeling experiments.

Abstract

The microbial type sesquiterpene synthase RlMTPSL4 from the liverwort Radula lindenbergiana was investigated for its products, showing the formation of several sesquiterpene hydrocarbons. The main product was structurally characterized as the new compound 4,5-diepi-isoishwarane, while the side products included the known hydrocarbons germacrene A, α-selinene, eremophilene and 4,5-diepi-aristolochene. The cyclization mechanism towards 4,5-diepi-isoishwarane catalyzed by RlMTPSL4 was investigated through isotopic labeling experiments, revealing the stereochemical course for the deprotonation step to the neutral intermediate germacrene A, a reprotonation for its further cyclization, and a 1,2-hydride shift along the cascade. The absolute configuration of 4,5-diepi-isoishwarane was determined using a stereoselective deuteration approach, revealing an absolute configuration typically observed for a microbial type sesquiterpene.