A Functional Switch Between Asperfumene and Fusicoccadiene Synthase and Entrance to Asperfumene Biosynthesis through a Vicinal Deprotonation-Reprotonation Process.

The diterpene synthase AfAS was identified from Aspergillus fumigatiaffinis. Its amino acid sequence and-according to a structural model-active site architecture are highly similar to those of the fusicocca-2,10(14)-diene synthase PaFS, but AfAS produces a structurally much more complex diterpene with a novel 6-5-5-5 tetracyclic skeleton called asperfumene. The cyclisation mechanism of AfAS was elucidated through isotopic labelling experiments and DFT calculations. The reaction cascade proceeds in its initial steps through similar intermediates as for the PaFS cascade, but then diverges through an unusual vicinal deprotonation-reprotonation process that triggers a skeletal rearrangement at the entrance to the steps leading to the unique asperfumene skeleton. The structural model revealed only one major difference between the active sites: The PaFS residue F65 is substituted by I65 in AfAS. Intriguingly, site-directed mutagenesis experiments with both diterpene synthases revealed that position 65 serves as a bidirectional functional switch for the biosynthesis of tetracyclic asperfumene versus structurally less complex diterpenes.

Absolute Configuration of Oxabornyl Polyenes Prugosenes A1-A3 and Structural Revision of Prugosene A2.

Oxabornyl polyenes represent a unique group of polyketides characterized by a central polyene core flanked by a conserved oxabornyl moiety and a structurally diverse oxygen heterocyclic ring. They are widely distributed in fungi and possess a variety of biological activities. Due to the significant spatial separation between the two stereogenic ring systems, it is difficult to establish their overall relative configurations. Here, we isolated three oxabornyl polyenes, prugosenes A1-A3 (1-3), from Talaromyces sp. JNU18266-01. Although these compounds were first reported from Penicillium rugulosum, their overall relative and absolute configurations remained unassigned. By employing ozonolysis in combination with ECD calculations, we were able to establish their absolute configurations, and additionally obtained seven new chemical derivatives (4-10). Notably, through NMR data analysis and quantum chemical calculations, we achieved the structural revision of prugosene A2. Furthermore, prugosenes A1-A3 exhibited potent antiviral activity against the respiratory syncytial virus, with compound 1 displaying an IC50 value of 6.3 µM. Our study thus provides a valuable reference for absolute configuration assignment of oxabornyl polyene compounds.