Discovery of non-squalene triterpenes

All known triterpenes are generated by triterpene synthases (TrTSs) from squalene or oxidosqualene1. This approach is fundamentally different from the biosynthesis of short-chain (C10–C25) terpenes that are formed from polyisoprenyl diphosphates2–4. In this study, two fungal chimeric class I TrTSs, Talaromyces verruculosustalaropentaene synthase (TvTS) and Macrophomina phaseolinamacrophomene synthase (MpMS), were characterized. Both enzymes use dimethylallyl diphosphate and isopentenyl diphosphate or hexaprenyl diphosphate as substrates, representing the first examples, to our knowledge, of non-squalene-dependent triterpene biosynthesis. The cyclization mechanisms of TvTS and MpMS and the absolute configurations of their products were investigated in isotopic labelling experiments. Structural analyses of the terpene cyclase domain of TvTS and full-length MpMS provide detailed insights into their catalytic mechanisms. An AlphaFold2-based screening platform was developed to mine a third TrTS, Colletotrichum gloeosporioidescolleterpenol synthase (CgCS). Our findings identify a new enzymatic mechanism for the biosynthesis of triterpenes and enhance understanding of terpene biosynthesis in nature.