A gene encoding a putative dimodular nonribosomal peptide synthetase (NRPS) was identified within a gene cluster of Aspergillus fumigatus, a species reported to produce fumitremorgins and other prenylated alkaloids. The gene was deleted and overexpressed in the genome reference strain Af293, and was also expressed in the naïve host Aspergillus nidulans, which lacks the equivalent gene cluster. While neither fumitremorgins nor the dipeptide brevianamide F (cyclo-L-Trp-L-Pro), an early intermediate, were detected in wild-type and deletion strains of A. fumigatus, brevianamide F accumulated in fungal cultures following increased expression of the NRPS gene in both A. fumigatus and A. nidulans. We conclude that the gene Afu8g00170, named ftmA, encodes the NRPS brevianamide synthetase. Brevianamide F is the precursor of a variety of fungal prenylated alkaloids with biological activity, including fumitremorgins A, B and C and tryprostatin B.

Biosynthesis of the flavonoid naringenin in plants and bacteria is commonly catalysed by a type III polyketide synthase (PKS) using one p-coumaroyl-CoA and three malonyl-CoA molecules as substrates. Here, we report a fungal non-ribosomal peptide synthetase -polyketide synthase (NRPS-PKS) hybrid FnsA for the naringenin formation. Feeding experiments with isotope-labelled precursors demonstrate that FnsA accepts not only p-coumaric acid (p-CA), but also p-hydroxybenzoic acid (p-HBA) as starter units, with three or four malonyl-CoA molecules for elongation, respectively. In vitro assays and MS/MS analysis prove that both p-CA and p-HBA are firstly activated by the adenylation domain of FnsA. Phylogenetic analysis reveals that the PKS portion of FnsA shares high sequence homology with type I PKSs. Refactoring the biosynthetic pathway in yeast with the involvement of fnsA provides an alternative approach for the production of flavonoids such as isorhamnetin and acacetin.

The following study depicts the development of a whole cell biocatalyst for the prenylation of indole derivatives. For this purpose the prenyltransferase FgaPT2 from Aspergillus fumigatus was displayed on the surface of Escherichia coli cells by using Autodisplay. The presence of the prenyltransferase in the outer membrane was detected by using SDS‐PAGE and Western Blot after the proteins of the outer membrane were isolated. The orientation of the prenyltransferase towards the outside of the cells was investigated by accessibility testing with externally added proteases. The FgaPT2 whole cell biocatalyst converted up to 250 μM of indole‐3‐propionic acid, approximately 25 % of the substrate used in the assay (100 μL sample, OD578=40). Another indole substrate, L‐β‐homotryptophan was also investigated and a conversion of 13 % was determined. By optimizing the assay conditions the conversion rate could be raised to approximately 30 % of indole‐3‐propionic acid during a 24 h incubation time at 20 °C. The whole cell biocatalyst endured a storage period of one month at 8 °C without any detectable loss in activity. Reusability was confirmed by recycling the biocatalyst. After three cycles of consecutive use, the whole cell biocatalyst retained a conversion rate of 46 % of indole‐3‐propionic acid and 23 % of L‐β‐homotryptophan after the third cycle.

The hydroxylated and diacetylated cyclo-l-Trp-l-Leu derivative (−)-protubonine B was isolated from a culture of Aspergillus ustus 3.3904. Genome mining led to the identification of a putative biosynthetic gene cluster coding for a bimodular nonribosomal peptide synthetase, a flavin-dependent monooxygenase, and two acetyltransferases. Heterologous expression of the pbo cluster in Aspergillus nidulans showed that it is responsible for the formation of the isolated metabolite. Gene deletion experiments and structural elucidation of the isolated intermediates confirmed the biosynthetic steps. In vitro experiments with the recombinant protein proved that the flavin-dependent oxygenase is responsible for stereospecific hydroxylation at the indole ring accompanied by pyrrolidine ring formation.

Annullatins from Cordyceps annullata are alkylated aromatic polyketides including annullatin D with a fused dihydrobenzofuran lactone ring system. Here, we report the identification of a silent biosynthetic gene cluster for annullatins from Penicillium roqueforti by heterologous expression in Aspergillus nidulans, gene deletion, and feeding experiments as well as by biochemical characterization. The polyketide core structure is consecutively modified by hydroxylation and prenylation. A berberine bridge enzyme-like protein catalyzes the final step, an oxidative lactonization between two hydroxyl groups, to form (2S, 9S)annullatin D.