Two Flavoenzymes Convert Fungal Polyketide-Nonribosomal Peptide to <i>cis</i> -2-Butene-1,4-dial-Type Carcinogen via Olefin Dihydroxylation and C–C Bond Cleavage

Postmodification steps by multiple oxidation enzymes are an effective strategy for converting the polyketide-nonribosomal peptide (PK-NRP) core backbone to structurally distinctive natural products, where decoration steps occurring on conjugated polyenes (olefin ≥3) of the PK-NRP core backbones are relatively scarce. In this study, we identified two flavoenzymes from the <i>frt</i> cluster in <i>Fusarium reticulatum</i>: (1) a FAD-dependent monooxygenase FrtD catalyzes olefin dihydroxylation of a five-conjugated polyene PK-NRP precursor (<b>8</b>) to yield vicinal diols (<b>1a</b>/<b>1b</b>); (2) a secretion-type berberine bridge enzyme-like oxidase FrtB is then responsible for converting <b>1a</b> to <i>cis-</i>2-methyl-2-butene-1,4-dial (<b>17</b>) and three conjugated polyene α,β-unsaturated ketone (<b>12</b>), the process of which includes dehydrogenation, C-C bond cleavage, and isomerization. Most importantly, <b>17</b> was characterized as a new <i>cis</i>-2-butene-1,4-dial-type carcinogen from <i>Fusarium</i> sp. due to its ability to bind amino acids and induce DNA damage. Our work reveals the unusual functions of flavoenzymes and discovers a new mycotoxin metabolite from <i>Fusarium</i> sp., which poses potential health risks that require further attention in the future.