title: Cloning and characterization of an Armillaria gallica cDNA encoding protoilludene synthase, which catalyzes the first committed step in the synthesis of antimicrobial melleolides.
abstract: Melleolides and related fungal sesquiterpenoid aryl esters are antimicrobial and cytotoxic natural products derived from cultures of the Homobasidiomycetes genus Armillaria. The initial step in the biosynthesis of all melleolides involves cyclization of the universal sesquiterpene precursor farnesyl diphosphate to produce protoilludene, a reaction catalyzed by protoilludene synthase. We achieved the partial purification of protoilludene synthase from a mycelial culture of Armillaria gallica and found that 6-protoilludene was its exclusive reaction product. Therefore, a further isomerization reaction is necessary to convert the 6-7 double bond into the 7-8 double bond found in melleolides. We expressed an A. gallica protoilludene synthase cDNA in Escherichia coli, and this also led to the exclusive production of 6-protoilludene. Sequence comparison of the isolated sesquiterpene synthase revealed a distant relationship to other fungal terpene synthases. The isolation of the genomic sequence identified the 6-protoilludene synthase to be present as a single copy gene in the genome of A. gallica, possessing an open reading frame interrupted with eight introns.
authors.affiliation: Fraunhofer Institut für Molekularbiologie und Angewandte Ökologie, Forckenbeckstrasse 6, 52074 Aachen, Germany.
authors.forename: Benedikt
authors.initials: B
authors.lastname: Engels
authors.forename: Uwe
authors.initials: U
authors.lastname: Heinig
authors.forename: Torsten
authors.initials: T
authors.lastname: Grothe
authors.forename: Marc
authors.initials: M
authors.lastname: Stadler
authors.forename: Stefan
authors.initials: S
authors.lastname: Jennewein
mesh_terms: D019883:Alkyl and Aryl Transferases / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D000890:Anti-Infective Agents / Q000378:metabolism*; D055392:Armillaria / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D015195:Drug Design; D004926:Escherichia coli / Q000235:genetics; D007438:Introns / Q000235:genetics; D008828:Microbiological Techniques; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D011106:Polyisoprenyl Phosphates / Q000378:metabolism; D012717:Sesquiterpenes / Q000737:chemistry / Q000378:metabolism*
chemical_list: D000890:Anti-Infective Agents; D001426:Bacterial Proteins; D018076:DNA, Complementary; D011106:Polyisoprenyl Phosphates; D012717:Sesquiterpenes; C004808:farnesyl pyrophosphate; D019883:Alkyl and Aryl Transferases; C108877:terpene synthase