Mechanistic investigations of hirsutene biosynthesis catalyzed by a chimeric sesquiterpene synthase from Steccherinum ochraceum

[Display omitted] •First heterologous expression of chimeric STS A8411 in Aspergillus oryzae.•The absolute configuration of hirsutene was determined by x-ray diffraction.•The Biosynthetic mechanism of hirsutene was elucidated by isotope labeling experiment. The high efficiency and elegance of terpene synthases is fascinating in constructing the molecular skeleton of complicated terpenoids with multiple chiral centers. Although the rapid development of sequencing technology has led to the discovery of an increasing number of terpene synthases, the cyclization mechanisms of some terpene synthases remains elusive. Here, we report that a chimeric sesquiterpene synthase from Steccherinum ochraceum is responsible for the biosynthesis of (+)-hirsutene, a linear triquinane sesquiterpene. Structural validation, and isotope labeling experiments demonstrate that the biosynthesis of (+)-hirsutene employs an unusual cyclization mode, involving three different cyclization processes (C1-C11, C2-C9, C3-C6), one intramolecular 1,2-hydride shift (C9-C10) and three successive 1,2-alkyl shifts to construct the 5–5–5 fused ring skeleton of (+)-hirsutene.