title: Elucidation of the biosynthesis of carnosic acid and its reconstitution in yeast.
abstract: Rosemary extracts containing the phenolic diterpenes carnosic acid and its derivative carnosol are approved food additives used in an increasingly wide range of products to enhance shelf-life, thanks to their high anti-oxidant activity. We describe here the elucidation of the complete biosynthetic pathway of carnosic acid and its reconstitution in yeast cells. Cytochrome P450 oxygenases (CYP76AH22-24) from Rosmarinus officinalis and Salvia fruticosa already characterized as ferruginol synthases are also able to produce 11-hydroxyferruginol. Modelling-based mutagenesis of three amino acids in the related ferruginol synthase (CYP76AH1) from S. miltiorrhiza is sufficient to convert it to a 11-hydroxyferruginol synthase (HFS). The three sequential C20 oxidations for the conversion of 11-hydroxyferruginol to carnosic acid are catalysed by the related CYP76AK6-8. The availability of the genes for the biosynthesis of carnosic acid opens opportunities for the metabolic engineering of phenolic diterpenes, a class of compounds with potent anti-oxidant, anti-inflammatory and anti-tumour activities.
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle 06120, Germany.
authors.forename: Ulschan
authors.initials: U
authors.lastname: Scheler
authors.affiliation: Department of Bioorganic Chemistry, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle 06120, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Brandt
authors.affiliation: Department of Bioorganic Chemistry, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle 06120, Germany.
authors.forename: Andrea
authors.identifier: http://orcid.org/0000-0002-8148-6895
authors.initials: A
authors.lastname: Porzel
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle 06120, Germany.
authors.forename: Kathleen
authors.initials: K
authors.lastname: Rothe
authors.affiliation: Program of Plant Metabolism and Metabolic Engineering, Centre for Research in Agricultural Genomics, Campus UAB, 08193 Bellaterra, Spain.|Faculty of Pharmacy, Department of Biochemistry and Molecular Biology, University of Barcelona, 08028 Barcelona, Spain.
authors.forename: David
authors.initials: D
authors.lastname: Manzano
authors.affiliation: Group of Biotechnology of Pharmaceutical Plants, Laboratory of Pharmacognosy, Department of Pharmaceutical Sciences, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.
authors.forename: Dragana
authors.initials: D
authors.lastname: Božić
authors.affiliation: Group of Biotechnology of Pharmaceutical Plants, Laboratory of Pharmacognosy, Department of Pharmaceutical Sciences, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.
authors.forename: Dimitra
authors.initials: D
authors.lastname: Papaefthimiou
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle 06120, Germany.
authors.forename: Gerd Ulrich
authors.initials: GU
authors.lastname: Balcke
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle 06120, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Henning
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle 06120, Germany.
authors.forename: Swanhild
authors.initials: S
authors.lastname: Lohse
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle 06120, Germany.
authors.forename: Sylvestre
authors.initials: S
authors.lastname: Marillonnet
authors.affiliation: Group of Biotechnology of Pharmaceutical Plants, Laboratory of Pharmacognosy, Department of Pharmaceutical Sciences, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.
authors.forename: Angelos K
authors.initials: AK
authors.lastname: Kanellis
authors.affiliation: Program of Plant Metabolism and Metabolic Engineering, Centre for Research in Agricultural Genomics, Campus UAB, 08193 Bellaterra, Spain.|Faculty of Pharmacy, Department of Biochemistry and Molecular Biology, University of Barcelona, 08028 Barcelona, Spain.
authors.forename: Albert
authors.initials: A
authors.lastname: Ferrer
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle 06120, Germany.
authors.forename: Alain
authors.identifier: http://orcid.org/0000-0002-9406-4245
authors.initials: A
authors.lastname: Tissier
mesh_terms: D045784:Abietanes / Q000096:biosynthesis* / Q000378:metabolism; D000893:Anti-Inflammatory Agents / Q000737:chemistry; D000970:Antineoplastic Agents / Q000737:chemistry; D000975:Antioxidants / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D016877:Oxidants / Q000737:chemistry; D010100:Oxygen; D010802:Phylogeny; D027542:Rosmarinus / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D027544:Salvia / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D045784:Abietanes; D000893:Anti-Inflammatory Agents; D000970:Antineoplastic Agents; D000975:Antioxidants; D016877:Oxidants; C431074:ferruginol; C068623:carnosol; C018381:salvin; D010100:Oxygen