The biosynthesis of the anti-microbial diterpenoid leubethanol inLeucophyllum frutescensproceeds via an all-cisprenyl intermediate

Serrulatane diterpenoids are natural products found in plants from a subset of genera within the figwort family (Scrophulariaceae). Many of these compounds have been characterized as having anti-microbial properties and share a common diterpene backbone. One example, leubethanol from Texas sage (Leucophyllum frutescens) has demonstrated activity against multi-drug-resistant tuberculosis. Leubethanol is the only serrulatane diterpenoid identified from this genus; however, a range of such compounds have been found throughout the closely relatedEremophilagenus. Despite their potential therapeutic relevance, the biosynthesis of serrulatane diterpenoids has not been previously reported. Here we leverage the simple product profile and high accumulation of leubethanol in the roots ofL.frutescensand compare tissue-specific transcriptomes with existing data fromEremophila serrulatato decipher the biosynthesis of leubethanol. A short-chaincis-prenyl transferase (LfCPT1) first produces the rare diterpene precursor nerylneryl diphosphate, which is cyclized by an unusual plastidial terpene synthase (LfTPS1) into the characteristic serrulatane diterpene backbone. Final conversion to leubethanol is catalyzed by a cytochrome P450 (CYP71D616) of the CYP71 clan. This pathway documents the presence of a short-chaincis-prenyl diphosphate synthase, previously only found in Solanaceae, which is likely involved in the biosynthesis of other known diterpene backbones inEremophila.LfTPS1 represents neofunctionalization of a compartment-switching terpene synthase accepting a novel substrate in the plastid. Biosynthetic access to leubethanol will enable pathway discovery to more complex serrulatane diterpenoids which share this common starting structure and provide a platform for the production and diversification of this class of promising anti-microbial therapeutics in heterologous systems.