title: A copal-8-ol diphosphate synthase from the angiosperm Cistus creticus subsp. creticus is a putative key enzyme for the formation of pharmacologically active, oxygen-containing labdane-type diterpenes.
abstract: The resin of Cistus creticus subsp. creticus, a plant native to Crete, is rich in labdane-type diterpenes with significant antimicrobial and cytotoxic activities. The full-length cDNA of a putative diterpene synthase was isolated from a C. creticus trichome cDNA library. The deduced amino acid sequence of this protein is highly similar (59%-70% identical) to type B diterpene synthases from other angiosperm species that catalyze a protonation-initiated cyclization. The affinity-purified recombinant Escherichia coli-expressed protein used geranylgeranyl diphosphate as substrate and catalyzed the formation of copal-8-ol diphosphate. This diterpene synthase, therefore, was named CcCLS (for C. creticus copal-8-ol diphosphate synthase). Copal-8-ol diphosphate is likely to be an intermediate in the biosynthesis of the oxygen-containing labdane-type diterpenes that are abundant in the resin of this plant. RNA gel-blot analysis revealed that CcCLS is preferentially expressed in the trichomes, with higher transcript levels found in glands on young leaves than on fully expanded leaves, while CcCLS transcript levels increased after mechanical wounding. Chemical analyses revealed that labdane-type diterpene production followed a similar pattern, with higher concentrations in trichomes of young leaves and increased accumulation upon wounding.
authors.affiliation: Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, Michigan 48109-1048, USA.
authors.forename: Vasiliki
authors.initials: V
authors.lastname: Falara
authors.forename: Eran
authors.initials: E
authors.lastname: Pichersky
authors.forename: Angelos K
authors.initials: AK
authors.lastname: Kanellis
mesh_terms: D019883:Alkyl and Aryl Transferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D055162:Biocatalysis; D029758:Cistus / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D018076:DNA, Complementary / Q000302:isolation & purification; D004224:Diterpenes / Q000737:chemistry / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D007700:Kinetics; D053858:Metabolic Networks and Pathways; D008969:Molecular Sequence Data; D010100:Oxygen / Q000378:metabolism*; D010802:Phylogeny; D018515:Plant Leaves / Q000235:genetics / Q000254:growth & development; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
chemical_list: D018076:DNA, Complementary; D004224:Diterpenes; D012333:RNA, Messenger; D011994:Recombinant Proteins; D019883:Alkyl and Aryl Transferases; D010100:Oxygen