MetaCyc Pathway: superpathway of pterocarpan biosynthesis (via formononetin)
Inferred from experiment

Pathway diagram: superpathway of pterocarpan biosynthesis (via formononetin)

If an enzyme name is shown in bold, there is experimental evidence for this enzymatic activity.

Superclasses: BiosynthesisSecondary Metabolites BiosynthesisPhenylpropanoid Derivatives BiosynthesisFlavonoids BiosynthesisIsoflavonoids Biosynthesis

Some taxa known to possess this pathway include : Cicer arietinum, Glycine max, Glycyrrhiza echinata, Glycyrrhiza glabra, Lotus japonicus, Medicago sativa, Medicago truncatula, Phaseolus vulgaris, Pisum sativum, Pueraria montana lobata

Expected Taxonomic Range: Fabaceae

General Background

Pterocarpans constitute a group of natural isoflavonoids, which are typically involved as phytoalexins in the defense against pathogens of leguminous plants. In contrast to many other isoflavonoids, pterocarpans are mostly not constitutively expressed but induced by biotic and abiotic stress [Dewick94].

Formononetin is a crucial intermediate in the biosynthesis of phytoalexins, and as such the starting point for the formation of various pterocarpans. The recent discovery of a new 4'-O-methyltransferase [Akashi03], which facilitates an additional biosynthetic route to synthesize formononetin, sheds new light on the intermediary steps of phytoalexin production in legumes.

The biosynthetic route of pterocarpans leads to various compounds distinctive for certain leguminous species. The simple pterocarpans medicarpin (alfalfa), maackiain (chickpea) and pisatin (pea) are characterized by methoxy, methylendioxy and 6a-hydroxyl ring substitutents. On the other side there are the complex type pterocarpans, such as phaseollins (bean) and gluceollins (soybean) bearing prenyl substituents on their furan ring structure [Dixon99] [Tahara].

About This Pathway

The biosynthesis of pterocarpans is salient by the stereo-specificity of many of the enzymes involved. Pterocarpan synthase, the enzyme converting 2'-hydroxyisoflavanones [e.g. (-)-vestitone] to the corresponding pterocarpans [e.g. (6aR, 11aR)-medicarpin] has been demonstrated in alfalfa to consist of in fact two enzymes, a reductase and a dehydratase [Guo94a] [Guo94]. The precedent enzyme, isoflavone reductase, catalyzes the stereo-specific formation of the 2'-hydroxyisoflavanones (6aS or 6aR configuration), which determines the stereochemistry of the subsequent pterocarpans, e.g. (+)-maackiain in pea and (-)-maackiain in chickpea.

Pterocarpan phytoalexins are antimicrobial compounds in leguminous plants formed upon pathogenic attacks and similar biotic stress-related events. Maackiain is together with medicarpin the main pterocarpan phytoalexin in chickpea and occurs there exclusively in a (-)-(6aR,11aR)-configuration [Guo94a]. Maackiain was allocated within the two subtribes Diocleinae and Erythrininae to 12 species representing 4 genera [Ingham90].

The biosynthetic route of maackiain starts with formononetin, a 4'-methoxylated isoflavone, which is also the origin for the pterocarpans medicarpin in chickpea and pisatin in pea. The biosynthetic route differs in essentially two additional steps, one introducing a hydroxyl group to the 3'-position of the isoflavone B-ring [Dewick78] followed by another one forming the methylendioxy bridge of maackiain [Clemens96].

The question of how the final stereochemistry is applied to the various pterocarpans is still under discussion. In leguminous plants pterocarpans have been found with different stereochemical orientation. Chick pea and alfalfa accumulate only (-)- or (6aR,11aR)-pterocarpans, whereas peanuts accumulate pterocarpans of the opposite stereochemistry (+)- (6aS, 11aS). The stereochemistry of this phytoalexins can be crucial since it has been found that pathogens cannot break down the opposite enantiomer of the pterocarpans usually found in their target plants [VanEtten83].

There are two possibilities to introduce stereo-specificity to pterocarpans, one is that the reduction of the first chiral center (6a), catalyzed by the isoflavanone reductase, determines the final stereochemistry of the pterocarpans. It has been argued that once this center is fixed the second chiral center (11a) cannot invert the stereochemistry which would cause undue ring strain [Guo94]. This would require isoflavone reductase(s) catalyzing two different stereospecific reactions.

The other possibility is that the stereochemistry alters after the first reduction. The isoflavones isolated to date possess only one configuration, which is the (-)- or (6aR,11aR). It is not clear yet how this configuration is converted into the opposite form such as in plants like pea and peanut, where the (+)-pterocarpans accumulate. It has been suggested that a special epimerase exists in this plants, which carries out the epimerization on the stage of either the isoflavanones or pterocarpanes [Dixon95]. However the existence of such an enzyme remains to be shown [Guo94a] [Guo94] [Guo95].

The detection and partial characterization of an new enzyme presumably involved in the formation of (+)-pisatin, i.e. isoflavene synthase indicates another path towards (+)-pisatin than the proposed isomerization of (-)-maackiain to (+)-maackiain. It has been previously demonstrated that the sophorol reductase catalyzed the reduction of (-)-sophorol and exclusively produced cis-(-)-7,2'-dihydroxy-4',5'-methylenedioxyisoflavanol and not its trans-enantiomer [DiCenzo06]. Only the former cis-enantiomer was shown to be accepted by the isoflavene synthase (IFSV) catalyzing the formation of 7,2'-dihydroxy-4',5'-methylenedioxyisoflav-3-ene from cis-(-)-7,2'-dihydroxy-4',5'-methylenedioxyisoflavanol via dehydrogenation [Celoy14]. This enzymatic reaction is only present in Pisum sativum known for producing the phytoalexin (+)-pisatin and does not operate in other legumes such as alfalfa, chickpea and bean which produce (-)-phytoalexins. Therefore it has been discussed that the reaction catalyzed by IFVS is a branching point towards pisatin biosynthesis although the step(s) forming (+)-enantiomeric intermediates still remain to be elucidated [Celoy14].

Subpathways: formononetin biosynthesis, medicarpin biosynthesis, maackiain biosynthesis

Created 13-Oct-2005 by Foerster H, TAIR
Revised 12-Feb-2015 by Foerster H, Boyce Thompson Institute


Akashi03: Akashi T, Sawada Y, Shimada N, Sakurai N, Aoki T, Ayabe S (2003). "cDNA cloning and biochemical characterization of S-adenosyl-L-methionine: 2,7,4'-trihydroxyisoflavanone 4'-O-methyltransferase, a critical enzyme of the legume isoflavonoid phytoalexin pathway." Plant Cell Physiol 44(2);103-12. PMID: 12610212

Celoy14: Celoy RM, VanEtten HD (2014). "(+)-Pisatin biosynthesis: from (-) enantiomeric intermediates via an achiral 7,2'-dihydroxy-4',5'-methylenedioxyisoflav-3-ene." Phytochemistry 98;120-7. PMID: 24332213

Clemens96: Clemens S, Barz W, (1996) "Cytochrome P450-dependent methylendioxy bridge formation in Cicer arietinum." Phytochemistry (1996) 41(2), 457-460.

Dewick78: Dewick PM, Ward D, (1978) "Isoflavone precursors of the pterocarpan phytoalexin maackiain in Trifolium pratense." Phytochemistry (1978) 17, 1751-1754.

Dewick94: Dewick PM (1994). "Isoflavonoids." In: Harborne JB (editor) The flavonoids: Advances in research since 1986; Chapman and Hall, London , 117-238.

DiCenzo06: DiCenzo GL, VanEtten HD (2006). "Studies on the late steps of (+) pisatin biosynthesis: evidence for (-) enantiomeric intermediates." Phytochemistry 67(7);675-83. PMID: 16504226

Dixon95: Dixon RA, Harrison MJ, Paiva NL, (1995) "The isoflavonoid phytoalexin pathway: From enzymes to genes to transcription factors." Physiologia Plantarum (1995), 93, 385-392.

Dixon99: Dixon RA, 1999 "Isoflavonoids: biochemistry, molecular biology, and biological functions." In: Comprehensive natural products chemistry Vol. 1: Sankawa, U. (editor), Polyketides and other secondary metabolites including fatty acids and their derivatives. Amsterdam, New York: Elsevier 1999, 773-82323.

Guo94: Guo L, Dixon RA, Paiva NL (1994). "The 'pterocarpan synthase' of alfalfa: association and co-induction of vestitone reductase and 7,2'-dihydroxy-4'-methoxy-isoflavanol (DMI) dehydratase, the two final enzymes in medicarpin biosynthesis." FEBS Lett 356(2-3);221-5. PMID: 7805842

Guo94a: Guo L, Dixon RA, Paiva NL (1994). "Conversion of vestitone to medicarpin in alfalfa (Medicago sativa L.) is catalyzed by two independent enzymes. Identification, purification, and characterization of vestitone reductase and 7,2'-dihydroxy-4'-methoxyisoflavanol dehydratase." J Biol Chem 269(35);22372-8. PMID: 8071365

Guo95: Guo L, Paiva NL (1995). "Molecular cloning and expression of alfalfa (Medicago sativa L.) vestitone reductase, the penultimate enzyme in medicarpin biosynthesis." Arch Biochem Biophys 320(2);353-60. PMID: 7625843

Ingham90: Ingham JL, (1990) "Systematic aspects of phytoalexin in formation within tribe Phaseoleae of the Leguminosae (subfamily Papilionoideae)." Biochemical Systematics and Ecology (1990), 18(5), 329-343.

Tahara: Tahara S, Ibrahim RK, (1995) "Prenylated isoflavonoids - an update." Phytochemistry, 38, 1073-1094.

VanEtten83: VanEtten HD, Matthews PS, Mercer EH, (1983) "(+)-maackiain and (+)-medicarpin as phytoalexins in Sophora japonica and identification of the (-) isomers by biotransformation." Phytochemistry (1983), 22(10), 2291-2295.

Other References Related to Enzymes, Genes, Subpathways, and Substrates of this Pathway

Akashi00: Akashi T, Sawada Y, Aoki T, Ayabe S (2000). "New scheme of the biosynthesis of formononetin involving 2,7,4'-trihydroxyisoflavanone but not daidzein as the methyl acceptor." Biosci Biotechnol Biochem 64(10);2276-9. PMID: 11129614

Akashi05: Akashi T, Aoki T, Ayabe S (2005). "Molecular and biochemical characterization of 2-hydroxyisoflavanone dehydratase. Involvement of carboxylesterase-like proteins in leguminous isoflavone biosynthesis." Plant Physiol 137(3);882-91. PMID: 15734910

Akashi06: Akashi T, VanEtten HD, Sawada Y, Wasmann CC, Uchiyama H, Ayabe S (2006). "Catalytic specificity of pea O-methyltransferases suggests gene duplication for (+)-pisatin biosynthesis." Phytochemistry 67(23);2525-30. PMID: 17067644

Akashi98: Akashi T, Aoki T, Ayabe S (1998). "CYP81E1, a cytochrome P450 cDNA of licorice (Glycyrrhiza echinata L.), encodes isoflavone 2'-hydroxylase." Biochem Biophys Res Commun 251(1);67-70. PMID: 9790908

Banks82: Banks SW, Dewick PM, (1982) "(-)-pisatin, an induced pterocarpan metabolite of abnormal configuration from Pisum sativum." Phytochemistry (1982), 21(7), 1605-1608.

Banks82a: Banks SW, Dewick PM, (1982) "Biosynthesis of the 6a-hydroxypterocarpan phytoalexin pisatin in Pisum sativum." Phytochemistry (1982), 21(9), 2235-2242.

Barz: Barz W, Welle R, (1992) "Biosynthesis and metabolism of isoflavones and pterocarpan phytoalexins in chickpea, soybean and phytopathogenic fungi." In: Stafford, H.A., Ibrahim, R.K. (editors) Phenolic metabolism in plants, recent advances in phytochemistry, Vol 26, Plenum Press New York and London, 139-164.

Barz89: Barz W, Bless W, Daniel S, Gunia W, Hinderer W, Jaques U, Kessmann H, Meier D, Tiemann K, Wittkampf U, (1989) "Elicitation and supression of isoflavones and pterocarpan phytoalexins in chickpea (Cicer arietinum L.) cell cultures." In: Kurz WGW (editor) Primary and secondary metabolism of plant cell cultures II, Springer Verlag Berlin Heidelberg New York London Paris Tokyo Hong Kong, (1989), 208-218.

Bauer91a: Bauer W, Zenk MH, (1991) "Two methylenedioxy bridge forming Cytochrome P-450 dependent enzymes are involved in (S)-stylopine biosynthesis." Phytochemistry (1991), 30(9), 2953-2961.

Bhandari92: Bhandari, P., Crombie, L., Daniels, P., Holden, I., Van Bruggen, N., Whiting, D. (1992). "Biosynthesis of the A/B/C/D-Ring System of the Rotenoid Amorphigeninby Amorpha fruticosa SeedIings." Journal of the Chemical Society, Perkin Transactions 1: 839 - 849.

Bless88: Bless W, Barz W, (1988) "Isolation of pterocarpan synthase, the terminal enzyme of pterocarpan phytoalexin biosynthesis in cell suspension cultures of Cicer arietinum." FEBS Letters (1988), 235(1), 47-50.

Clemens: Clemens S, Hinderer W, Wittkampg U, Barz W, (1993) "Characterization of cytochrome P450-dependent isoflavone hydroxylase from chickpea." Phytochemistry, 32(3), 653-657.

Crombie98: Crombie L, Whiting DA (1998). "Review article number 135 biosynthesis in the rotenoid group of natural products: applications of isotope methodology." Phytochemistry 49(6);1479-1507. PMID: 11711058

Daniel90: Daniel S, Tiemann K, Wittkampf U, Bless W, Hinderer W, Barz W, (1990) "Elicitor-induced metabolic changes in cell cultures of chickpea (Cicer arietinum L.) cultivar resistant and susceptible to Ascochyta rabiei." Planta (1990), 182(2), 270-278.

George98: George HL, Hirschi KD, VanEtten HD (1998). "Biochemical properties of the products of cytochrome P450 genes (PDA) encoding pisatin demethylase activity in nectria haematococca." Arch Microbiol 170(3);147-54. PMID: 9683653

Gunia: Gunia W, Hinderer W, Wittkampf U, Barz W, (1991) "Elicitor induction of cytochrome P-450 monooxygenases in cell suspension cultures of chickpea (Cicer arietinum L.) and their involvement in pterocarpan phytoalexin biosynthesis." Z. Naturforsch. 46c, 58-66.

Hagmann84: Hagmann ML, Heller W, Grisebach H (1984). "Induction of phytoalexin synthesis in soybean. Stereospecific 3,9-dihydroxypterocarpan 6a-hydroxylase from elicitor-induced soybean cell cultures." Eur J Biochem 142(1);127-31. PMID: 6540173

Hagmann84a: Hagmann M, Grisebach H, (1984) "Enzymatic rearrangement of flavanone to isoflavone." FEBS Letters (1984), 175(2), 199-202.

Hakamatsuka98: Hakamatsuka T, Mori K, Ishida S, Ebizuka Y, Sankawa U (1998). "Purification of 2-hydroxyisoflavanone dehydratase from the cell cultures of Pueraria lobata." Phytochemistry 49(2);497-505.

Heller94: Heller W, Forkmann G (1994). "Biosynthesis of flavonoids." In: Harborne JB (editor) The flavonoids. Advances in research since 1986. Chapman & Hall, London Glasgow New York Tokyo Melbourne Madras, 499-537.

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Please cite the following article in publications resulting from the use of MetaCyc: Caspi et al, Nucleic Acids Research 42:D459-D471 2014
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