Arabidopsis thaliana isoprenyl diphosphate synthases produce the C25 intermediate geranylfarnesyl diphosphate.
Isoprenyl diphosphate synthases (IDSs) catalyze some of the most basic steps in terpene biosynthesis by producing the prenyl diphosphate precursors of each of the various terpenoid classes. Most plants investigated have distinct enzymes that produce the short-chain all-trans (E) prenyl diphosphates geranyl diphosphate (GDP, C10 ), farnesyl diphosphate (FDP, C15 ) or geranylgeranyl diphosphate (GGDP, C20 ). In the genome of Arabidopsis thaliana, 15 trans-product-forming IDSs are present. Ten of these have recently been shown to produce GGDP by genetic complementation of a carotenoid pathway engineered into Escherichia coli. When verifying the product pattern of IDSs producing GGDP by a new LC-MS/MS procedure, we found that five of these IDSs produce geranylfarnesyl diphosphate (GFDP, C25 ) instead of GGDP as their major product in enzyme assays performed in vitro. Over-expression of one of the GFDP synthases in A. thaliana confirmed the production of GFDP in vivo. Enzyme assays with A. thaliana protein extracts from roots but not other organs showed formation of GFDP. Furthermore, GFDP itself was detected in root extracts. Subcellular localization studies in leaves indicated that four of the GFDP synthases were targeted to the plastoglobules of the chloroplast and one was targeted to the mitochondria. Sequence comparison and mutational studies showed that the size of the R group of the 5th amino acid residue N-terminal to the first aspartate-rich motif is responsible for C25 versus C20 product formation, with smaller R groups (Ala and Ser) resulting in GGDP (C20 ) as a product and a larger R group (Met) resulting in GFDP (C25 ).
Department of Biochemistry, Max Planck Institute for Chemical Ecology, Beutenberg Campus, Hans Knoell Straße 8, D-07745 Jena, Germany.
Raimund
R
Nagel
Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany.
Carolin
C
Bernholz
Institute of Biology and Ecology, Pavol Jozef Šafárik University Košice, Mánesova 23, 04154 Košice, Slovakia.
Eva
E
Vranová
Institute of Biology and Ecology, Pavol Jozef Šafárik University Košice, Mánesova 23, 04154 Košice, Slovakia.
Ján
J
Košuth
Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany.
Nick
N
Bergau
Department of Bioorganic Chemistry, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany.
Steve
S
Ludwig
Department of Bioorganic Chemistry, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany.
Ludger
L
Wessjohann
Department of Biochemistry, Max Planck Institute for Chemical Ecology, Beutenberg Campus, Hans Knoell Straße 8, D-07745 Jena, Germany.
Jonathan
J
Gershenzon
Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany.
Alain
A
Tissier
Department of Biochemistry, Max Planck Institute for Chemical Ecology, Beutenberg Campus, Hans Knoell Straße 8, D-07745 Jena, Germany.
Axel
A
Schmidt
D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000032:analysis / Q000737:chemistry / Q000502:physiology*; D004926:Escherichia coli / Q000235:genetics; D051231:Farnesyltranstransferase / Q000032:analysis / Q000737:chemistry / Q000502:physiology*; D051228:Geranyltranstransferase / Q000032:analysis / Q000737:chemistry / Q000502:physiology*; D053858:Metabolic Networks and Pathways; D008928:Mitochondria / Q000378:metabolism; D008969:Molecular Sequence Data; D018087:Plastids / Q000378:metabolism; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein
D029681:Arabidopsis Proteins; D051228:Geranyltranstransferase; D051231:Farnesyltranstransferase
Arabidopsis thaliana; geranylfarnesyl diphosphate; geranylgeranyl diphosphate; isoprenyl diphosphate synthase; prenyltransferase; sesterterpenes; terpenes