Comparative characterization of fungal anthracenone and naphthacenedione biosynthetic pathways reveals an α-hydroxylation-dependent Claisen-like cyclization catalyzed by a dimanganese thioesterase.

摘要: The linear tetracyclic TAN-1612 (1) and BMS-192548 (2) were isolated from different filamentous fungal strains have been examined as potential neuropeptide Y neurokinin-1 receptor antagonists, respectively. Although the biosynthesis of aromatic polyketides has attracted much interest in recent years, biosynthetic mechanism for such naphthacenedione-containing products not established. Using a targeted genome mining approach, we first located ada gene cluster responsible 1 Aspergillus niger ATCC 1015. connection between pathway was verified through overexpression Zn(2)Cys(6)-type pathway-specific transcriptional regulator AdaR subsequent expression analysis. enzymes encoded share high sequence similarities to known apt linked anthraquinone asperthecin 3. Subsequent comparative investigation these two highly homologous clusters by heterologous reconstitution Saccharomyces cerevisiae revealed novel α-hydroxylation-dependent Claisen cyclization cascade, which involves flavin-dependent monooxygenase that hydroxylates α-carbon an acyl carrier protein-bound polyketide bifunctional metallo-β-lactamase-type thioesterase (MβL-TE). MβL-TE catalyzes fourth ring afford naphthacenedione scaffold upon α-hydroxylation, whereas it performs hydrolytic release anthracenone product absence α-hydroxylation. Through vitro biochemical characterizations metal analyses, is dimanganese enzyme requires both Mn(2+) cations observed activities. example Claisen-like condensation without α/β hydrolase fold forms no covalent bond with substrate. These mechanistic features should be general compounds fungi.