Biosynthesis of furanocoumarins: mevalonate-independent prenylation of umbelliferone in Apium graveolens (Apiaceae)

Stanjek, Volker; Piel, Jörn; Boland, Wilhelm

doi:10.1016/s0031-9422(98)00650-5

Cited by 50 publications

(32 citation statements)

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“…A primer was designed corresponding to the amino acid sequence Arg 27 -Leu 33 of Cyp73A1 fused to residues Pro 38 -Pro 45 of Cyp71AJ1 (reverse primer 5Ј-GGGATAT-TGTGGTGGAGAAGGTGGGAGCTTGAATTTTTTACCG-CGG) and employed for PCR with the CYP73A1 cDNA template in combination with another primer (forward 5Ј-GGATCCATGGACCTCCTCCTCATAGA) creating a BamHI restriction site upstream of the ATG. This first amplification generated a product of 123 nt composed of the upstream 99 nt of CYP73A1 followed downstream by 24 nt of CYP71AJ1.…”

Section: Methodsmentioning

confidence: 99%

“…1). Whereas the subcellular compartment and the precise mode of cyclization of 4-coumaric acid to umbelliferone have remained elusive (31), the 6-prenyltransferase was assigned to plastids in Ruta graveolens (32), which is compatible with the mevalonate-independent prenylation of umbelliferone observed in Apium graveolens (33). An equivalent location is likely for the 8-prenyltransferase activity, although this has not been as firmly documented.…”

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confidence: 90%

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Molecular Cloning and Functional Characterization of Psoralen Synthase, the First Committed Monooxygenase of Furanocoumarin Biosynthesis

Larbat

Kellner

Specker

et al. 2007

Journal of Biological Chemistry

135

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Ammi majus L. accumulates linear furanocoumarins by cytochrome P450 (CYP)-dependent conversion of 6-prenylumbelliferone via (؉)-marmesin to psoralen. Relevant activities, i.e. psoralen synthase, are induced rapidly from negligible background levels upon elicitation of A. majus cultures with transient maxima at 9 -10 h and were recovered in labile microsomes. Expressed sequence tags were cloned from elicited Ammi cells by a nested DD-RT-PCR strategy with CYP-specific primers, and full-size cDNAs were generated from those fragments correlated in abundance with the induction profile of furanocoumarin-specific activities. One of these cDNAs representing a transcript of maximal abundance at 4 h of elicitation was assigned CYP71AJ1. Functional expression in Escherichia coli or yeast cells initially failed but was accomplished eventually in yeast cells after swapping the N-terminal membrane anchor domain with that of CYP73A1. The recombinant enzyme was identified as psoralen synthase with narrow substrate specificity for (؉)-marmesin. Psoralen synthase catalyzes a unique carbon-chain cleavage reaction concomitantly releasing acetone by syn-elimination. Related plants, i.e. Heracleum mantegazzianum, are known to produce both linear and angular furanocoumarins by analogous conversion of 8-prenylumbelliferone via (؉)-columbianetin to angelicin, and it was suggested that angelicin synthase has evolved from psoralen synthase. However, (؉)-columbianetin failed as substrate but competitively inhibited psoralen synthase activity. Analogy modeling and docked solutions defined the conditions for high affinity substrate binding and predicted the minimal requirements to accommodate (؉)-columbianetin in the active site cavity. The studies suggested that several point mutations are necessary to pave the road toward angelicin synthase evolution.Furanocoumarins are produced by many plants, mostly of the Apiaceae, Rutaceae, Moraceae, or the Coronilla and Psoralea genera of the Fabaceae (1-3). Multiple pharmacological effects have been ascribed to several of these metabolites (4 -6), which were included in clinical screenings but received attention also for their inhibitory effect on monooxygenases involved in drug metabolism (7-9) and potential toxicity (10). The (dihydro)furan-substituted 2H-1-benzopyran-2-one forms the characteristic core structure, and the annulation type distinguishes the linear furanocoumarins or psoralens from the angular furanocoumarins (Fig.

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Section: Methodsmentioning

confidence: 99%

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confidence: 90%

Molecular Cloning and Functional Characterization of Psoralen Synthase, the First Committed Monooxygenase of Furanocoumarin Biosynthesis

Larbat

Kellner

Specker

et al. 2007

Journal of Biological Chemistry

135

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“…Previous studies using stereospecifically deuterated (ϩ)-marmesin confirmed that psoralen synthase activity catalyzes a single concerted reaction, initiated by the abstraction of the C-3Ј-hydrogen synconfigured to the isopropyloxy group and release of acetone as a by-product ( Fig. 1) (26,27). An equivalent study demonstrated by feeding leaves of Heracleum mantegazzianum with stereospecifically 3Ј-deuterated (ϩ)-columbianetin that the conversion of (ϩ)-columbianetin to angelicin (i.e.…”

mentioning

confidence: 88%

Isolation and Functional Characterization of CYP71AJ4 Encoding for the First P450 Monooxygenase of Angular Furanocoumarin Biosynthesis

Larbat¹,

Hehn²,

Hans

et al. 2009

Journal of Biological Chemistry

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The biosynthesis of linear and angular furanocoumarins is still poorly understood at the molecular level, with only psoralen synthase (CYP71AJ1) identified from Ammi majus. Using cDNA probes inferred from CYP71AJ1, three orthologs were isolated from Apium graveolens (CYP71AJ2) and Pastinaca sativa (CYP71AJ3 and -4) and functionally expressed in yeast cells. CYP71AJ2 and CYP71AJ3 displayed psoralen synthase activity, whereas CYP71AJ4 only catalyzed the conversion of (؉)-columbianetin to angelicin and negligible amounts of a hydroxylated columbianetin by-product. CYP71AJ4 thus constitutes the first fully characterized P450 monooxygenase specific for the angular furanocoumarin pathway. The angelicin synthase exhibited an apparent K m of 2.1 ؎ 0.4 M for (؉)-columbianetin and a k cat of 112 ؎ 14 min ؊1 . Moreover, the use of 3-deuterated (؉)-columbianetin as substrate led to an almost complete "metabolic switch," resulting in the synthesis of anti-3-hydroxy-3-deuterated(؉)-columbianetin. This confirms that angelicin synthase attacks columbianetin by syn-elimination of hydrogen from C-3. Sequence comparison between psoralen synthase (CYP71AJ3) and angelicin synthase (CYP71AJ4) showed 70% identity, whereas the identity dropped to 40% in those regions thought to provide the substrate recognition sites. Accordingly, CYP71AJ3 and CYP71AJ4 might be derived from a common ancestor of unknown functionality by gene duplication and subsequent molecular evolution.Furanocoumarins are natural plant metabolites characterized by a furane moiety fused to benzopyran-2-one. The position of the furane substitution distinguishes two large groups of compounds, the linear (psoralens) and the angular furanocoumarins (angelicin and derivatives) (Fig.

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“…The pathway to the phosphorylated hemiterpenes is located either in plastids (DOX pathway) or in the cytosol (MVA pathway). Both pathways are involved in the biosynthesis of different terpenoid natural products, but sesquiterpenes are an exception in that their terpenoid skeletons may be derived from either the DOX or the MVA pathway or simultaneously from both (Piel et al, 1998;Stanjek et al, 1999;Laule et al, 2003;Page et al, 2004;Dudareva et al, 2005;Flü gge and Gao, 2005). During the incubation experiment discussed here, we have not only analyzed ergoline alkaloids for a possible isotopic deuterium enrichment, but also (E)-b-caryophyllene, a sesquiterpene known to be a product of the I. asarifolia plant but not the associated fungus IasaF13 (Kucht et al, 2004 Figure 3.…”

Section: Presence Of Genes Responsible For Ergoline Alkaloid Biosynthmentioning

confidence: 99%

Biosynthesis and Accumulation of Ergoline Alkaloids in a Mutualistic Association between Ipomoea asarifolia (Convolvulaceae) and a Clavicipitalean Fungus

et al. 2008

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Biosynthesis of furanocoumarins: mevalonate-independent prenylation of umbelliferone in Apium graveolens (Apiaceae)

Cited by 50 publications

References 19 publications

Molecular Cloning and Functional Characterization of Psoralen Synthase, the First Committed Monooxygenase of Furanocoumarin Biosynthesis

Molecular Cloning and Functional Characterization of Psoralen Synthase, the First Committed Monooxygenase of Furanocoumarin Biosynthesis

Isolation and Functional Characterization of CYP71AJ4 Encoding for the First P450 Monooxygenase of Angular Furanocoumarin Biosynthesis

Biosynthesis and Accumulation of Ergoline Alkaloids in a Mutualistic Association between Ipomoea asarifolia (Convolvulaceae) and a Clavicipitalean Fungus

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