Construction of high efficient bio-production systems using filamentous fungi as host organism

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Construction of biological production system using Aspergillus oryzae as a host was investigated. Newly cloned polyketide synthase (PKS) gene, pksN, from Alternaria solani, was expressed under a-amylase promoter. The A. oryzae transformant produced new polyketide compound named alternapyrone, which has 8 methyl groups derived from methionine on the decaketide carbon backbone. To analyze the function of naphthopyrone synthase WA, modified derivatives of WA PKS were expressed. As a result, the C-terminus region which was so far considered to be thioesterase motif was found to catalyze Claisen-type cyclization involved in cyclization/aromatization of polyketomethylene chain. By the co-expression experiment of alb1 and ayg1 genes from Aspergillus fumigatus, it was found that alb1 gene encodes PKS for naphthopyrone YWA1 and ayg1 encodes for the enzyme Ayglp which converts YWA1 to 1, 3, 6, 8-tetrahydroxynaphthalene. Ayglp was overexpressed, purified, and characterized. To examine the expression of plant biosynthetic enzyme, expression plasmid for oxidosqualene cyclase was constructed and introduced into A. oryzae. However, expression of oxidosqalene cyclase was not detected.