Diterpene synthases of the biosynthetic system of medicinally active diterpenoids in Marrubium vulgare. (23rd July 2014)

• Record Type: Journal Article
• Authors: Zerbe, Philipp; Chiang, Angela; Dullat, Harpreet; O'Neil‐Johnson, Mark; Starks, Courtney; Hamberger, Björn; Bohlmann, Jörg
• Is Part Of: Plant journal. Volume 79:Number 6(2014:Sep.)
• Page Start: 914
• Page End: 927
• Publication Date: 2014-07-23
• Subjects: Plant molecular biology -- Periodicals; Plant cells and tissues -- Periodicals; Botany -- Periodicals; 580
• DOI: 10.1111/tpj.12589
• Languages: English
• ISSNs: 0960-7412

Abstract

<abstract abstract-type="main" id="tpj12589-abs-0001"> <title>Summary</title> <p> <italic>Marrubium vulgare</italic> (Lamiaceae) is a medicinal plant whose major bioactive compounds, marrubiin and other labdane–related furanoid diterpenoids, have potential applications as anti–diabetics, analgesics or vasorelaxants. Metabolite and transcriptome profiling of <italic>M. vulgare</italic> leaves identified five different candidate diterpene synthases (diTPSs) of the TPS–c and TPS–e/f clades. We describe the <italic>in vitro</italic> and <italic>in vivo</italic> functional characterization of the <italic>M. vulgare</italic> diTPS family. In addition to <italic>Mv</italic>EKS <italic>ent</italic>–kaurene synthase of general metabolism, we identified three diTPSs of specialized metabolism: <italic>Mv</italic>CPS3 (+)‐copalyl diphosphate synthase, and the functional diTPS pair <italic>Mv</italic>CPS1 and <italic>Mv</italic>ELS. In a sequential reaction, <italic>Mv</italic>CPS1 and <italic>Mv</italic>ELS produce a unique oxygenated diterpene scaffold 9, 13‐epoxy‐labd‐14‐ene en route to marrubiin and an array of related compounds. In contrast with previously known diTPSs that introduce a hydroxyl group at carbon C–8 of the labdane backbone, the <italic>Mv</italic>CPS1‐catalyzed reaction proceeds via oxygenation of an intermediate carbocation at C–9, yielding the bicyclic peregrinol diphosphate. <italic>Mv</italic>ELS belongs to a subgroup of the diTPS TPS–e/f clade with unusual βα–domain architecture. <italic>Mv</italic>ELS is active <italic>in vitro</italic> and <italic>in vivo</italic> with three different prenyl diphosphate substrates forming the marrubiin precursor 9, 13‐epoxy‐labd‐14‐ene, as identified by nuclear magnetic resonance (NMR) analysis, manoyl oxide and miltiradiene. <italic>Mv</italic>ELS fills a central position in the biosynthetic system that forms the foundation for the diverse repertoire of <italic>Marrubium</italic> diterpenoids. Co‐expression of <italic>Mv</italic>CPS1 and <italic>Mv</italic>ELS in engineered <italic>E. coli</italic> and <italic>Nicotiana benthamiana</italic> offers opportunities for producing precursors for an array of biologically active diterpenoids.</p> </abstract>