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Highlights • A target fishing protocol was developed to define activity of orphan food compounds. • The fungal secondary metabolite atromentin served for the proof of concept. • A combined in silico/in vitro approach succeeded in activity identification. • Estrogen receptors and 17-β-HSDs were identified as relevant biological targets. • Mild estrogenicity of atromentin was experimentally confirmed. Abstract Many small molecules of food origin may effect human health but lack an adequate description of their biological activity. To fill this knowledge gap, a first-line workflow is needed to assign putative functions, rank the endpoints for testing and guide wet-lab experiments. In this framework, the identification of potential biological targets can be used to probe the activity of orphan compounds using a so-called “target fishing” approach. Here, we present a proof of concept study using an in silico/in vitro target fishing approach on the fungal secondary metabolite atromentin. The procedure relies on a computational screening for activity identification coupled with experimental trials for dose-response characterization. Computational results identified estrogen receptors and 17-β-hydroxysteroid dehydrogenase as potential targets. Experiments confirmed a weak estrogenic activity, supporting the reliability of the procedure. Despite limited estrogenicity of atromentin, the proposed inhibition of 17-β-hydroxysteroid dehydrogenase should be considered as a source for endocrine disruptive effects. [ABSTRACT FROM AUTHOR]

Abstract: This report highlights the first biochemical characterization of a multi-domain biosynthetic enzyme for basidiomycete secondary metabolism: the tri-domain enzyme atromentin synthetase AtrA, from Tapinella panuoides, which adenylates and dimerizes 4-hydroxyphenylpyruvic acid into atromentin. Also, the l-tyrosine:2-oxoglutarate aminotransferase AtrD, which provides the substrate for this dimerization step, has been characterized. AtrA and AtrD expand the shikimic acid pathway from l-tyrosine to atromentin, the central terphenylquinone intermediate for a prominent and widely occurring class of basidiomycete pigments, among them various pharmaceutically relevant compounds. The genes atrA and atrD were cloned and found to be clustered within one genetic locus. Given the broad distribution of atromentin-derived compounds among homobasidiomycetes we expect our system represents a widely applicable model. [Copyright &y& Elsevier]

The article offers information regarding the establishment of the structure of atromentin, a long-known natural pigment, through the single-crystal X-ray analysis and total synthesis based on double Suzuki-Miyauro coupling. It presents the process of the total synthesis which started with Suzuki-Miyaura coupling and the use of oxidation. It adds that the preparation of 2-o-methoxyatromentin and thelepantin was done through the application of ceric ammonium nitrate (CAN) oxidation.

Summary Non-ribosomal peptide synthetase (NRPS)-like enzymes catalyze the non-oxidative homodimerization of aromatic α-keto acids, but the exact reaction mechanism is unknown. The furanone-forming thioesterase domain of the Aspergillus terreus aspulvinone E synthetase MelA displays a predicted quinone-forming motif, whereby its catalytic triad contains an essential cysteine indicating an unusual thioester intermediate. To convert MelA into a quinone-forming atromentin synthetase its thioesterase domain was replaced with that from a Paxillus involutus or A. terreus atromentin synthetase. Phylogenetic proximity of donor and acceptor seems important, as only replacement with the A. terreus thioesterase was functional. Heterologous expression of atromentin synthetases in Aspergillus niger and Aspergillus oryzae revealed host-dependent product formation whereby cross-chemistry directed atromentin biosynthesis in A. niger toward atrofuranic acid. Screening of aspergilli from section Nigri identified an atromentin synthetase in Aspergillus brasiliensis that produced atrofuranic acid in the homologous host. Therefore, cross-chemistry on quinone cores appears common to section Nigri. Graphical Abstract Highlights • The product spectrum of NRPS-like enzymes depends on the expression platform • Cross-chemistry occurs on quinone core structures in aspergilli from section Nigri • Cross-chemistry produces novel metabolites such as atrofuranic acid • Identification of an atromentin synthetase from Aspergillus brasiliensis Geib et al. investigated core structure formation by thioesterase domains of fungal NRPS-like enzymes. Exchange of thioesterase domains alters product formation, but requires close phylogenetic relationship of donor and acceptor. Product formation further depends on expression hosts with furanic acid rather than quinone core formation in aspergilli from section Nigri. [ABSTRACT FROM AUTHOR]