Pentaketide melanin biosynthesis in Aspergillus fumigatus requires chain-length shortening of a heptaketide precursor

Chain lengths and cyclization patterns of microbial polyketides are general ly determined by polyketide synthases alone. Fungal polyketide melanins are often derived from a pentaketide 1,8-dihydroxynaphthalene, and pentaketide synthases are used for synthesis of the upstream pentaketide precursor, 1, 3,6,8-tetrahydroxynaphthalene (1,3,6,8-THN). However, Aspergillus fumigatus , a human fungal pathogen, uses a heptaketide synthase (Alb1p) to synthesiz e its conidial pigment through a pentaketide pathway similar to that which produces 1,8-dihydroxynaphthalene-melanin. In this study we demonstrate tha t a novel protein, Ayg1p, is involved in the formation of 1,3,6,8-THN by ch ain-length shortening of a heptaketide precursor in A. fumigatus. Deletion of the ayg1 gene prevented the accumulation of 1,3,6,8-THN suggesting the i nvolvement of ayg1 in 1,3,6,8-THN production. Genetic analyses of double-ge ne deletants suggested that Ayg1p catalyzes a novel biosynthetic step downs tream of Alb1p and upstream of Arp2p (1,3,6,8-THN reductase). Further genet ic and biochemical analyses of the reconstituted strains carrying alb1, ayg 1, or alb1 + ayg1 indicated that Ayg1p is essential for synthesis of 1,3,6, 8-THN in addition to Alb1p. Cell-free enzyme assays, using the crude Ayg1p protein extract, revealed that Ayg1p enzymatically shortened the heptaketid e product of Alb1p to 1,3,6,8-THN. Thus, the protein Ayg1p facilitates the participation of a heptaketide synthase in a pentaketide pathway via a nove l polyketide-shortening mechanism in A fumigatus.