Reconstitution of biosynthetic pathway for mushroom-derived cyathane diterpenes in yeast and generation of new "non-natural" analogues.

Mushroom-derived cyathane-type diterpenes possess unusual chemical skeleton and diverse bioactivities. To efficiently supply bioactive cyathanes for deep studies and explore their structural diversity, synthesis of cyathane diterpenes in a geranylgeranyl pyrophosphate engineered is investigated. Aided by homologous analyses, one new unclustered FAD-dependent oxidase EriM accounting for the formation of allyl aldehyde and three new NADP(H)-dependent reductases in the biosynthesis of cyathanes are identified and elucidated. By combinatorial biosynthetic strategy, strains generating twenty-two cyathane-type diterpenes, including seven "unnatural" cyathane xylosides (, , , , , , and ) are established. Compounds -, , and show significant neurotrophic effects on PC12 cells in the dose of 6.3-25.0 μmol/L. These studies provide new insights into the divergent biosynthesis of mushroom-originated cyathanes and a straightforward approach to produce bioactive cyathane-type diterpenes.