Cytochrome P450s from Cynara cardunculus L. CYP71AV9 and CYP71BL5, catalyze distinct hydroxylations in the sesquiterpene lactone biosynthetic pathway.

Abstract

Cynara cardunculus (Asteraceae) is a cross pollinated perennial crop which includes the two cultivated taxa globe artichoke and cultivated cardoon. The leaves of these plants contain high concentrations of sesquiterpene lactones (STLs) among which cynaropicrin is the most represented, and has recently attracted attention because of its therapeutic potential as anti-tumor and anti-photoaging agent. Costunolide is considered the common precursor of the STLs and three enzymes are involved in its biosynthetic pathway: i.e. the germacrene A synthase (GAS), the germacrene A oxidase (GAO) and the costunolide synthase (COS). Here we report on the isolation of two P450 genes, (i.e. CYP71AV9 and CYP71BL5), in a set of ∼19,000 C. cardunculus unigenes, and their functional characterization in yeast and in planta. The metabolite analyses revealed that the co-expression of CYP71AV9 together with GAS resulted in the biosynthesis of germacra-1(10),4,11(13)-trien-12-oic acid in yeast. The co-expression of CYP71BL5 and CYP71AV9 with GAS led to biosynthesis of the free costunolide in yeast and costunolide conjugates in Nicotiana benthamiana, demonstrating their involvement in STL biosynthesis as GAO and COS enzymes. The substrate specificity of CYP71AV9 was investigated by testing its ability to convert amorpha-4,11-diene, (+)-germacrene D and cascarilladiene to their oxidized products when co-expressed in yeast with the corresponding terpene synthases.

• Authors: Kaouthar Eljounaidi, Katarina Cankar, Cinzia Comino, Andrea Moglia, Alain Hehn, Frédéric Bourgaud, Harro Bouwmeester, Barbara Menin, Sergio Lanteri, Jules Beekwilder
• Journal: Plant science : an international journal of experimental plant biology (Plant Sci) Vol. 223 Pg. 59-68 (Jun 2014) ISSN: 1873-2259 [Electronic] Ireland
• PMID: 24767116 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Chemical References

• Lactones
• Plant Proteins
• Sesquiterpenes
• Sesquiterpenes, Germacrane
• germacrene A
• costunolide
• Cytochrome P-450 Enzyme System
• Oxidoreductases
• Alkyl and Aryl Transferases
• terpene synthase

Topics

• Alkyl and Aryl Transferases (metabolism)
• Biocatalysis
• Biosynthetic Pathways
• Cloning, Molecular
• Cynara (enzymology, genetics)
• Cytochrome P-450 Enzyme System (metabolism)
• Genes, Plant
• Hydroxylation
• Lactones (chemistry, metabolism)
• Mass Spectrometry
• Oxidoreductases (metabolism)
• Phylogeny
• Plant Proteins (metabolism)
• Plants, Genetically Modified
• Saccharomyces cerevisiae (metabolism)
• Sequence Alignment
• Sesquiterpenes (chemistry, metabolism)
• Sesquiterpenes, Germacrane (biosynthesis, chemistry)
• Substrate Specificity
• Tobacco (genetics)