Publications

• Chrysanthemyl diphosphate synthase operates in planta as a bifunctional enzyme with chrysanthemol synthase activity.

  Yang T., Gao L., Hu H., Stoopen G., Wang C., Jongsma M.A.

  Chrysanthemyl diphosphate synthase (CDS) is the first pathway-specific enzyme in the biosynthesis of pyrethrins, the most widely used plant-derived pesticide. CDS catalyzes c1'-2-3 cyclopropanation reactions of two molecules of dimethylallyl diphosphate (DMAPP) to yield chrysanthemyl diphosphate ( ... >> More

  Chrysanthemyl diphosphate synthase (CDS) is the first pathway-specific enzyme in the biosynthesis of pyrethrins, the most widely used plant-derived pesticide. CDS catalyzes c1'-2-3 cyclopropanation reactions of two molecules of dimethylallyl diphosphate (DMAPP) to yield chrysanthemyl diphosphate (CPP). Three proteins are known to catalyze this cyclopropanation reaction of terpene precursors. Two of them, phytoene and squalene synthase, are bifunctional enzymes with both prenyltransferase and terpene synthase activity. CDS, the other member, has been reported to perform only the prenyltransferase step. Here we show that the NDXXD catalytic motif of CDS, under the lower substrate conditions prevalent in plants, also catalyzes the next step, converting CPP into chrysanthemol by hydrolyzing the diphosphate moiety. The enzymatic hydrolysis reaction followed conventional Michaelis-Menten kinetics, with a Km value for CPP of 196 μm. For the chrysanthemol synthase activity, DMAPP competed with CPP as substrate. The DMAPP concentration required for half-maximal activity to produce chrysanthemol was ∼100 μm, and significant substrate inhibition was observed at elevated DMAPP concentrations. The N-terminal peptide of CDS was identified as a plastid-targeting peptide. Transgenic tobacco plants overexpressing CDS emitted chrysanthemol at a rate of 0.12-0.16 μg h(-1) g(-1) fresh weight. We propose that CDS should be renamed a chrysanthemol synthase utilizing DMAPP as substrate. << Less

  J. Biol. Chem. 289:36325-36335(2014) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.

• The seco-iridoid pathway from Catharanthus roseus.

  Miettinen K., Dong L., Navrot N., Schneider T., Burlat V., Pollier J., Woittiez L., van der Krol S., Lugan R., Ilc T., Verpoorte R., Oksman-Caldentey K.M., Martinoia E., Bouwmeester H., Goossens A., Memelink J., Werck-Reichhart D.

  The (seco)iridoids and their derivatives, the monoterpenoid indole alkaloids (MIAs), form two large families of plant-derived bioactive compounds with a wide spectrum of high-value pharmacological and insect-repellent activities. Vinblastine and vincristine, MIAs used as anticancer drugs, are prod ... >> More

  The (seco)iridoids and their derivatives, the monoterpenoid indole alkaloids (MIAs), form two large families of plant-derived bioactive compounds with a wide spectrum of high-value pharmacological and insect-repellent activities. Vinblastine and vincristine, MIAs used as anticancer drugs, are produced by Catharanthus roseus in extremely low levels, leading to high market prices and poor availability. Their biotechnological production is hampered by the fragmentary knowledge of their biosynthesis. Here we report the discovery of the last four missing steps of the (seco)iridoid biosynthesis pathway. Expression of the eight genes encoding this pathway, together with two genes boosting precursor formation and two downstream alkaloid biosynthesis genes, in an alternative plant host, allows the heterologous production of the complex MIA strictosidine. This confirms the functionality of all enzymes of the pathway and highlights their utility for synthetic biology programmes towards a sustainable biotechnological production of valuable (seco)iridoids and alkaloids with pharmaceutical and agricultural applications. << Less

  Nat. Commun. 5:3606-3606(2014) [PubMed] [EuropePMC]

  This publication is cited by 10 other entries.

• Coexpression analysis identifies two oxidoreductases involved in the biosynthesis of the monoterpene acid moiety of natural pyrethrin insecticides in Tanacetum cinerariifolium.

  Xu H., Moghe G.D., Wiegert-Rininger K., Schilmiller A.L., Barry C.S., Last R.L., Pichersky E.

  Flowers of <i>Tanacetum cinerariifolium</i> produce a set of compounds known collectively as pyrethrins, which are commercially important pesticides that are strongly toxic to flying insects but not to most vertebrates. A pyrethrin molecule is an ester consisting of either trans-chrysanthemic acid ... >> More

  Flowers of <i>Tanacetum cinerariifolium</i> produce a set of compounds known collectively as pyrethrins, which are commercially important pesticides that are strongly toxic to flying insects but not to most vertebrates. A pyrethrin molecule is an ester consisting of either trans-chrysanthemic acid or its modified form, pyrethric acid, and one of three alcohols, jasmolone, pyrethrolone, and cinerolone, that appear to be derived from jasmonic acid. Chrysanthemyl diphosphate synthase (CDS), the first enzyme involved in the synthesis of trans-chrysanthemic acid, was characterized previously and its gene isolated. <i>TcCDS</i> produces free trans-chrysanthemol in addition to trans-chrysanthemyl diphosphate, but the enzymes responsible for the conversion of trans-chrysanthemol to the corresponding aldehyde and then to the acid have not been reported. We used an RNA sequencing-based approach and coexpression correlation analysis to identify several candidate genes encoding putative trans-chrysanthemol and trans-chrysanthemal dehydrogenases. We functionally characterized the proteins encoded by these genes using a combination of in vitro biochemical assays and heterologous expression in planta to demonstrate that <i>TcADH2</i> encodes an enzyme that oxidizes trans-chrysanthemol to trans-chrysanthemal, while <i>TcALDH1</i> encodes an enzyme that oxidizes trans-chrysanthemal into trans-chrysanthemic acid. Transient coexpression of <i>TcADH2</i> and <i>TcALDH1</i> together with <i>TcCDS</i> in <i>Nicotiana benthamiana</i> leaves results in the production of trans-chrysanthemic acid as well as several other side products. The majority (58%) of trans-chrysanthemic acid was glycosylated or otherwise modified. Overall, these data identify key steps in the biosynthesis of pyrethrins and demonstrate the feasibility of metabolic engineering to produce components of these defense compounds in a heterologous host. << Less

  Plant Physiol. 176:524-537(2018) [PubMed] [EuropePMC]

  This publication is cited by 15 other entries.