(+)-Germacrene A biosynthesis: the committed step in the biosynthesis of bitter sesquiterpene lactones in chicory

• Published in: Plant physiology (Bethesda) 1998-08, Vol.117 (4), p.1381-1392
• Main Authors: Kraker, J.W. de (Wageningen Agricultural University, Wageningen, The Netherlands.), Franssen, M.C.R, Groot, A. de, Konig, W.A, Bouwmeester, H.J
• Format: Article
• Language: English
• Subjects: ACTIVIDAD ENZIMATICA; ACTIVITE ENZYMATIQUE; Agronomy. Soil science and plant productions; AMARGO; AMERTUME; Biochemistry and Enzymology; Biological and medical sciences; BIOSINTESIS; BIOSYNTHESE; BIOSYNTHESIS; BITTERNESS; CHEMICAL COMPOSITION; Chemical constitution; CHEMICAL STRUCTURE; CICHORIUM INTYBUS; COMPOSICION QUIMICA; COMPOSITION CHIMIQUE; Economic plant physiology; Enzyme activity; Enzymes; ENZYMIC ACTIVITY; ESTRUCTURA QUIMICA; Ethers; FEUILLE; Fundamental and applied biological sciences. Psychology; Gels; HOJAS; Instituut voor Agrobiologisch en Bodemvruchtbaarheidsonderzoek; Laboratorium voor Organische chemie; Laboratory for Organic Chemistry; LACTONAS; LACTONE; LACTONES; LEAVES; LIGASAS; LIGASE; LIGASES; MOLECULAR WEIGHT; Organic Chemistry; Organische Chemie; Pentanes; PESO MOLECULAR; POIDS MOLECULAIRE; RACINE; RAICES; Research Institute for Agrobiology and Soil Fertility; ROOTS; Sesquiterpenes; SESQUITERPENOID LACTONES; SESQUITERPENOIDE; SESQUITERPENOIDS; SESQUITERPENOS; Skeleton; STRUCTURE CHIMIQUE; Terpenes; VLAG
• ISSN: 0032-0889; 1532-2548
• DOI: 10.1104/pp.117.4.1381

The leaves and especially the roots of chicory (Cichorium intybus L.) contain high concentrations of bitter sesquiterpene lactones such as the guianolides lactupicrin, lactucin, and 8-deoxylactucin. Eudesmanolides and germacranolides are present in smaller amounts. Their postulated biosynthesis through the mevalonatefarnesyl diphosphate-germacradiene pathway has now been confirmed by the isolation of a (+)-germacrene A synthase from chicory roots. This sesquiterpene cyclase was purified 200-fold using a combination of anion-exchange and dye-ligand chromatography. It has a K(m) value of 6.6 micromolar, an estimated molecular mass of 54 kD, and a (broad) pH optimum around 6.7. Germacrene A, the enzymatic product, proved to be much more stable than reported in literature. Its heat-induced Cope rearrangement into (-)-beta-elemene was utilized to determine its absolute configuration on an enantio-selective gas chromatography column. To our knowledge, until now in sesquiterpene biosynthesis, germacrene A has only been reported as an (postulated) enzyme-bound intermediate, which, instead of being released, is subjected to additional cyclization(s) by the same enzyme that generated it from farnesyl diphosphate. However, in chicory germacrene A is released from the sesquiterpene cyclase. Apparently, subsequent oxidations and/or glucosylation of the germacrane skeleton, together with a germacrene cyclase, determine whether guaiane- or eudesmane-type sesquiterpene lactones are produced