Regiospecific hydroxylation of isoflavones by cytochrome P450 81E enzymes from <i>Medicago truncatula</i>

Liu, Changjun; Huhman, David V.; Sumner, Lloyd W.; Dixon, Richard A.

doi:10.1046/j.1365-313x.2003.01893.x

Cited by 138 publications

(92 citation statements)

References 51 publications

(100 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…1 are potentially suspect; for example, the products of 4CL-like genes may not use 4-coumarate as substrate (28,29). Likewise, of those genes upstream of formononetin that are not induced by MJ, recombinant TC100787 does not possess isoflavone 2Ј-hydroxylase activity despite its annotation (30), and not all isoflavone reductase (IFR)-like genes encode proteins with IFR activity (31).…”

Section: Discussionmentioning

confidence: 99%

Different mechanisms for phytoalexin induction by pathogen and wound signals in Medicago truncatula

Naoumkina

Farag

Sumner

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

171

157

View full text Add to dashboard Cite

Cell suspensions of the model legume Medicago truncatula accumulated the isoflavonoid phytoalexin medicarpin in response to yeast elicitor or methyl jasmonate (MJ), accompanied by decreased levels of isoflavone glycosides in MJ-treated cells. DNA microarray analysis revealed rapid, massive induction of early (iso)flavonoid pathway gene transcripts in response to yeast elicitor, but not MJ, and differential induction by the two elicitors of sets of genes encoding transcription factors, ABC transporters, and ␤-glucosidases. In contrast, both elicitors induced genes encoding enzymes for conversion of the isoflavone formononetin to medicarpin. Four MJ-induced ␤-glucosidases were expressed as recombinant enzymes in yeast, and three were active with isoflavone glucosides. The most highly induced ␤-glucosidase was nuclear localized and preferred flavones to isoflavones. The results indicate that the genetic and biochemical mechanisms underlying accumulation of medicarpin differ depending on the nature of the stimulus and suggest a role for MJ as a signal for rapid hydrolysis of preformed, conjugated intermediates for antimicrobial biosynthesis during wound responses.glucosidase ͉ methyl jasmonate ͉ phytoanticipin ͉ cell culture ͉ elicitation

show abstract

Section: Discussionmentioning

confidence: 99%

Different mechanisms for phytoalexin induction by pathogen and wound signals in Medicago truncatula

Naoumkina

Farag

Sumner

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

171

157

View full text Add to dashboard Cite

show abstract

“…The destination vectors were transformed into yeast strain W303A1 using the modified LiCl method (Pompon et al, 1996;Liu et al, 2003), and the transformants were selected on SD (2uracil) plates. Colonies confirmed by colony PCR were used for enzyme assay.…”

Section: Construction Of Yeast Expression Vector and Yeast Transformamentioning

confidence: 99%

Characterization of an Isoflavonoid-Specific Prenyltransferase from Lupinus albus

et al. 2012

Self Cite

View full text Add to dashboard Cite

Prenylated flavonoids and isoflavonoids possess antimicrobial activity against fungal pathogens of plants. However, only a few plant flavonoid and isoflavonoid prenyltransferase genes have been identified to date. In this study, an isoflavonoid prenyltransferase gene, designated as LaPT1, was identified from white lupin (Lupinus albus). The deduced protein sequence of LaPT1 shared high homologies with known flavonoid and isoflavonoid prenyltransferases. The LaPT1 gene was mainly expressed in roots, a major site for constitutive accumulation of prenylated isoflavones in white lupin. LaPT1 is predicted to be a membrane-bound protein with nine transmembrane regions and conserved functional domains similar to other flavonoid and isoflavonoid prenyltransferases; it has a predicted chloroplast transit peptide and is plastid localized. A microsomal fraction containing recombinant LaPT1 prenylated the isoflavone genistein at the B-ring 39 position to produce isowighteone. The enzyme is also active with 29-hydroxygenistein but has no activity with other flavonoid substrates. The apparent K m of recombinant LaPT1 for the dimethylallyl diphosphate prenyl donor is in a similar range to that of other flavonoid prenyltransferases, but the apparent catalytic efficiency with genistein is considerably higher. Removal of the transit peptide increased the apparent overall activity but also increased the K m . Medicago truncatula hairy roots expressing LaPT1 accumulated isowighteone, a compound that is not naturally produced in this species, indicating a strategy for metabolic engineering of novel antimicrobial compounds in legumes.

show abstract

“…Yeast cells were collected and resuspended in 0.1 M potassium phosphate, pH 8.0, containing 0.4 M Suc, 14 mM b-mercaptoethanol, and 13 protease inhibitors (Complete EDTA-free Protease Inhibitor Cocktail tablets; Roche Diagnostics). Microsomes were isolated as described (Liu et al, 2003). The microsomal pellet was resuspended in 0.1 M potassium phosphate, pH 8.0, containing 0.4 M Suc and 0.5 mM reduced glutathione.…”

Section: Biosynthesis and Purification Of 2749-trihydroxyisoflavanonementioning

confidence: 99%

Structural Basis for Dual Functionality of Isoflavonoid O-Methyltransferases in the Evolution of Plant Defense Responses

et al. 2006

Self Cite

View full text Add to dashboard Cite

In leguminous plants such as pea (Pisum sativum), alfalfa (Medicago sativa), barrel medic (Medicago truncatula), and chickpea (Cicer arietinum), 49-O-methylation of isoflavonoid natural products occurs early in the biosynthesis of defense chemicals known as phytoalexins. However, among these four species, only pea catalyzes 3-O-methylation that converts the pterocarpanoid isoflavonoid 6a-hydroxymaackiain to pisatin. In pea, pisatin is important for chemical resistance to the pathogenic fungus Nectria hematococca. While barrel medic does not biosynthesize 6a-hydroxymaackiain, when cell suspension cultures are fed 6a-hydroxymaackiain, they accumulate pisatin. In vitro, hydroxyisoflavanone 49-O-methyltransferase (HI49OMT) from barrel medic exhibits nearly identical steady state kinetic parameters for the 49-O-methylation of the isoflavonoid intermediate 2,7,49-trihydroxyisoflavanone and for the 3-O-methylation of the 6a-hydroxymaackiain isoflavonoid-derived pterocarpanoid intermediate found in pea. Protein x-ray crystal structures of HI49OMT substrate complexes revealed identically bound conformations for the 2S,3R-stereoisomer of 2,7,49-trihydroxyisoflavanone and the 6aR,11aR-stereoisomer of 6a-hydroxymaackiain. These results suggest how similar conformations intrinsic to seemingly distinct chemical substrates allowed leguminous plants to use homologous enzymes for two different biosynthetic reactions. The three-dimensional similarity of natural small molecules represents one explanation for how plants may rapidly recruit enzymes for new biosynthetic reactions in response to changing physiological and ecological pressures.

show abstract

Regiospecific hydroxylation of isoflavones by cytochrome P450 81E enzymes from Medicago truncatula

Cited by 138 publications

References 51 publications

Different mechanisms for phytoalexin induction by pathogen and wound signals in Medicago truncatula

Different mechanisms for phytoalexin induction by pathogen and wound signals in Medicago truncatula

Characterization of an Isoflavonoid-Specific Prenyltransferase from Lupinus albus

Structural Basis for Dual Functionality of Isoflavonoid O-Methyltransferases in the Evolution of Plant Defense Responses

Contact Info

Product

Resources

About