title: Total biosynthesis of diterpene aphidicolin, a specific inhibitor of DNA polymerase α: heterologous expression of four biosynthetic genes in Aspergillus oryzae.
abstract: Clustering of biosynthetic genes for producing fungal secondary metabolites, which frequently consist of less than ten genes, has been recognized with numerous genomes. The heterologous expression of whole genes in the clusters will therefore produce various types of natural products when using a suitable fungal host. We introduced the whole gene cluster for the biosynthesis of diterpene aphidicolin into the fungal quadruple auxotrophic host, Aspergillus oryzae, by using four different vectors (pTAex3, pPTRI, pUSA and pAdeA) which harbor a starch-inducible promoter/terminator to examine the expression conditions. The resulting quadruple transformant carrying the genes of geranylgeranyl diphosphate synthase PbGGS, terpene synthase PbACS, and two monooxygenases (PbP450-1 and PbP450-2) produced aphidicolin. The double and triple transformants also respectively produced aphidicolan-16β-ol and 3-deoxyaphidicolin. Alternative host Saccharomyces cerevisiae carrying the genes, PbGGS and PbACS, produced key intermediate aphidicolan-16β-ol. This is the first example of a total biosynthesis of terpenoids using fungal hosts.
authors: Division of Chemistry, Graduate School of Science, Hokkaido University, Sapporo.
authors: Ryuya
authors: R
authors: Fujii
authors: Atsushi
authors: A
authors: Minami
authors: Tae
authors: T
authors: Tsukagoshi
authors: Natsuko
authors: N
authors: Sato
authors: Takehiko
authors: T
authors: Sahara
authors: Satoru
authors: S
authors: Ohgiya
authors: Katsuya
authors: K
authors: Gomi
authors: Hideaki
authors: H
authors: Oikawa
mesh_terms: D019883:Alkyl and Aryl Transferases / Q000235:genetics / Q000378:metabolism*; D016590:Aphidicolin / Q000096:biosynthesis*; D001236:Aspergillus oryzae / Q000201:enzymology / Q000235:genetics*; D004256:DNA Polymerase I / Q000037:antagonists & inhibitors; D004224:Diterpenes / Q000737:chemistry / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism* / Q000494:pharmacology; D051231:Farnesyltranstransferase / Q000235:genetics / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D005800:Genes, Fungal; D005818:Genetic Engineering / Q000379:methods; D005822:Genetic Vectors; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism*; D005810:Multigene Family; D011401:Promoter Regions, Genetic; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D013729:Terpenes / Q000737:chemistry / Q000378:metabolism*; D014170:Transformation, Genetic
chemical_list: D004224:Diterpenes; D004791:Enzyme Inhibitors; D013729:Terpenes; D016590:Aphidicolin; C037612:3-deoxyaphidicolin; D006899:Mixed Function Oxygenases; D019883:Alkyl and Aryl Transferases; C108877:terpene synthase; D051231:Farnesyltranstransferase; D004256:DNA Polymerase I