Abstract

Diketomorpholines (DKMs) represent an intriguing class of biologically active heterocycles. To identify unusual DKM natural products, we conducted a comprehensive analysis of fungal genomes from NCBI and our laboratory databases. A non-ribosomal peptide synthetase (NRPS) biosynthetic gene cluster (BGC) was identified in Aspergillus aculeatus CRI323-04, which also harbors a dimethylallyl tryptophan synthase (DMATS) gene and an NAD(P)H-dependent reductase gene. Heterologous expression of this BGC in chassis organism A. nidulans resulted in the production of prenylated DKM (3), seco-diketomorpholines (seco-DKMs) with regular (6–7) or reverse prenyl group (8), as well as rare ketooxadiazepane alkaloids (4–5). Through integrated in vivo and in vitro experiments, we elucidated the biosynthetic pathway of the prenylated DKM (3). Moreover, the feeding experiment showed that the complex conversion of prenylated DKM (3) to seco-DKMs (6–8) and ketooxadiazepane (4–5) alkaloids may be mediated by heterologous host A. nidulans. This study elucidates the biosynthetic pathway of 3 and further facilitates the exploration of structurally diverse prenylated DKMs and ketooxadiazepane alkaloids from fungal resources.