Tolypocladamide H and the Proposed Tolypocladamide NRPS in Species.

The genome of entomopathogenic fungus Gams encodes 43 putative biosynthetic gene clusters for specialized metabolites, although genotype-phenotype linkages have been reported only for the cyclosporins and fumonisins. was cultured in defined minimal media, supplemented with or without one of nine different amino acids. Acquisition of LC-MS/MS data for molecular networking and manual analysis facilitated annotation of putative known and unknown metabolites. These data led us to target a family of peptaibols and guided the isolation and purification of tolypocladamide H (), which showed modest antibacterial activity and toxicity to mammalian cells at micromolar concentrations. HRMS/MS, NMR, and advanced Marfey's analysis were used to assign the structure of as a peptaibol containing 4-[()-2-butenyl]-4-methyl-l-threonine (Bmt), a hallmark structural motif of the cyclosporins. LC-MS detection of homologous tolypocladamide metabolites and phylogenomic analyses of peptaibol biosynthetic genes in other cultured species allowed assignment of a putative tolypocladamide nonribosomal peptide synthetase gene.