Structural evidence for the inhibition of grape dihydroflavonol 4-reductase by flavonols

Trabelsi, Nadia; Petit, P.; Manigand, Claude; d’Estaintot, Béatrice Langlois; Granier, T.; Chaudière, Jean; Gallois, B.

doi:10.1107/s0907444908017769

Cited by 25 publications

(24 citation statements)

References 35 publications

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“…This is well documented in proline-rich proteins which are believed to play an important role in the astringency of polyphenols [33]. In Vv-DFR, we even described the 3D-structure of a crystallised quaternary complex containing NADPH and a pair of stacking molecules of quercetin [34]. Interestingly, in leucoanthocyanidin reductase from Vitis vinifera [35], another member of the short-chain dehydrogenase family whose substrate is the product of DFR, the productive binding configuration of the tricyclic structure is head-to-tail with respect to that observed in the homologous ternary abortive complex of Vv-DFR [15].…”

Section: Discussionmentioning

confidence: 99%

Kinetic and binding equilibrium studies of dihydroflavonol 4-reductase from Vitis vinifera and its unusually strong substrate inhibition

Trabelsi¹,

d’Estaintot²,

Sigaud³

et al. 2011

JBPC

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Dihydroflavonol 4-reductase (DFR), a member of the short-chain dehydrogenase family, catalyzes the last common step in the biosynthesis of flavan-3-ols and condensed tannins. Initial rates of DFR were measured by monitoring the 340-nm absorbance decrease resulting from the joint consumption of dihydroquercetin (DHQ) and NADPH, as a function of pH, temperature and ionic strength. At pH 6.5 and 30˚C, substrate inhibition was observed above 30 µM DHQ. At lower/non-inhibitory DHQ concentrations, NADP + behaves as a competitive inhibitor with respect to NADPH and as a mixed inhibitor with respect to DHQ, which supports a sequential ordered mechanism, with NADPH binding first and NADP + released last. Binding-equilibrium data obtained by means of the chromatographic method of Hummel and Dreyer at pH 7.5 and by isothermal calorimetric titration at pH 6.5 led to the conclusion that ligands of the apoenzyme included NADPH, NADP + and DHQ. The mechanism which best accounts for substrate inhibition at pH 6.5 in the absence of product involves the formation of a binary non-productive E.DHQ complex. Thus, a productive ternary complex cannot be formed when DHQ binds first. This mechanism of inhibition may prevent the accumulation of unstable leucoanthocyanidins within cells.

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Section: Discussionmentioning

confidence: 99%

Kinetic and binding equilibrium studies of dihydroflavonol 4-reductase from Vitis vinifera and its unusually strong substrate inhibition

Trabelsi¹,

d’Estaintot²,

Sigaud³

et al. 2011

JBPC

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“…Although it was not the highest matches in our Dali and BLAST searches, dihydroflavonol reductase (DFR) from V. vinifera, which catalyzes the NADPHdependent reduction of dihydroquercetin, is the only one that offers the ternary complex structure with its substrate (Trabelsi et al, 2008). Given that this dihydroflavonol and coniferaldehyde share critical aspects of chemical architecture, and that SbCCR1 contains functional analogs of DFR substrate-binding residues, we deemed the DFR an appropriate model for comparisons.…”

Section: Discussion Active Site Kinetics Of Enzyme Reaction and Catmentioning

confidence: 99%

Structural and Biochemical Characterization of Cinnamoyl-CoA Reductases

et al. 2016

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Cinnamoyl-coenzyme A reductase (CCR) catalyzes the reduction of hydroxycinnamoyl-coenzyme A (CoA) esters using NADPH to produce hydroxycinnamyl aldehyde precursors in lignin synthesis. The catalytic mechanism and substrate specificity of cinnamoyl-CoA reductases from sorghum (Sorghum bicolor), a strategic plant for bioenergy production, were deduced from crystal structures, site-directed mutagenesis, and kinetic and thermodynamic analyses. Although SbCCR1 displayed higher affinity for caffeoyl-CoA or p-coumaroyl-CoA than for feruloyl-CoA, the enzyme showed significantly higher activity for the latter substrate. Through molecular docking and comparisons between the crystal structures of the Vitis vinifera dihydroflavonol reductase and SbCCR1, residues threonine-154 and tyrosine-310 were pinpointed as being involved in binding CoA-conjugated phenylpropanoids. Threonine-154 of SbCCR1 and other CCRs likely confers strong substrate specificity for feruloyl-CoA over other cinnamoyl-CoA thioesters, and the T154Y mutation in SbCCR1 led to broader substrate specificity and faster turnover. Through data mining using our structural and biochemical information, four additional putative CCR genes were discovered from sorghum genomic data. One of these, SbCCR2, displayed greater activity toward p-coumaroyl-CoA than did SbCCR1, which could imply a role in the synthesis of defense-related lignin. Taken together, these findings provide knowledge about critical residues and substrate preference among CCRs and provide, to our knowledge, the first three-dimensional structure information for a CCR from a monocot species.

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“…The stacking of the flavonoids myricetin and quercetin in the active site may inhibit enzymatic activity. "The X-ray structures of both DFR-NADP (+)-myricetin and DFR-NADP (+)-quercetin have been deposited and published together with preliminary spectroscopic data" [29]. The structural complexes demonstrate that "flavonols could be inhibitors of the activity of DFR towards dihydroflavonols" in a feedback inhibition mechanism [29].…”

Section: Hydroxy Containing Flavonoids (Quercetin) Inhibit Xanthine Omentioning

confidence: 99%

“…Structural complexes of DFR and flavonoids myricetin and quercetin showed significant changes with respect to that of the previously reported DFR-NADP (+)-DHQ complex. "Two flavonol molecules bind to the catalytic site in a stacking arrangement and alter its geometry" [29]. The stacking of the flavonoids myricetin and quercetin in the active site may inhibit enzymatic activity.…”

Section: Hydroxy Containing Flavonoids (Quercetin) Inhibit Xanthine Omentioning

confidence: 99%

“…The leucoanthocyanidins are the precursors of anthocyanins and condensed tannins, two major classes of phenolic compounds that strongly influence the bioorganic medicinal properties of wine. DFR has been investigated in many plant species, structural properties were unknown until the three-dimensional structure of the Vitis vinifera enzyme complexed with NADP (+) and its natural substrate dihydroquercetin (DHQ) were solved" [29]. Crystals of DFR-NADP (+)-flavonol (myricetin and quercetin) complexes were obtained and their structures solved in order to better understand substrate specificity.…”

Section: Hydroxy Containing Flavonoids (Quercetin) Inhibit Xanthine Omentioning

confidence: 99%

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Structural Analysis of Flavonoid/Drug Target Complexes: Natural Products as Lead Compounds for Drug Development

Sosa¹,

Sosa²,

Phansalkar³

et al. 2017

Nat Prod Chem Res

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Plants produce pigments called flavonoids that are synthesized (de novo) primarily from the amino acid phenylalanine. Many natural products such as herb preparations for medicinal use contain flavonoids and so these diverse structural compounds have become potential lead compounds for a variety of illnesses ranging from specific cancers to gout. Recently, Biotechnology has been utilized through bioorganic engineering to manufacture flavonoid and modified flavonoids for medicinal applications. A growing number of protein-flavonoid complexes have been crystallized and their structures solved in the last decade. A summary of the protein-flavonoid complexes with medicinal relevance is presented herein. A detailed analysis of selected protein-flavonoid complexes is provided. The goal is to provide insights for modifications to known flavonoids that could be biomanufactured to generate more specific and efficient binding of flavonoids to protein targets with medicinal relevance.

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Structural evidence for the inhibition of grape dihydroflavonol 4-reductase by flavonols

Cited by 25 publications

References 35 publications

Kinetic and binding equilibrium studies of dihydroflavonol 4-reductase from Vitis vinifera and its unusually strong substrate inhibition

Kinetic and binding equilibrium studies of dihydroflavonol 4-reductase from Vitis vinifera and its unusually strong substrate inhibition

Structural and Biochemical Characterization of Cinnamoyl-CoA Reductases

Structural Analysis of Flavonoid/Drug Target Complexes: Natural Products as Lead Compounds for Drug Development

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