1.13.11.24: quercetin 2,3-dioxygenase
This is an abbreviated version!
For detailed information about quercetin 2,3-dioxygenase, go to the full flat file.
Word Map on EC 1.13.11.24
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1.13.11.24
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flavonols
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dioxygenation
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bicupins
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o-heterocycle
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oxygenolysis
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synthesis
- 1.13.11.24
- flavonols
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dioxygenation
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bicupins
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o-heterocycle
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oxygenolysis
- synthesis
Reaction
Synonyms
2,3-QD, 2,3QD, 2,4-QD, Co-QDO, Co-QueD, Cu2+-containing 2,4-QD, cupin domain-containing protein, Fe-QDO, Fe-QueD, flavonol 2,4-dioxygenase, flavonol 2,4-oxygenase, manganese quercetin 2,3-dioxygenase, manganese quercetin dioxygenase, Mn-QDO, Mn-QueD, Ni-QueD, nickel quercetinase, pirin, QDO, QdoI, QueD, quercetin 2,4-dioxygenase, quercetin dioxygenase, quercetinase, type III extradiol dioxygenase, VdQase, YxaG
ECTree
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Metals Ions
Metals Ions on EC 1.13.11.24 - quercetin 2,3-dioxygenase
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Co2+
copper
Cu
Cu2+
Fe2+
HNO
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nitrosyl hydride replaces dioxygen in nitroxygenase activity of manganese quercetin dioxygenase resulting in the incorporation of both N and O atoms into the product. Turnover is demonstrated by consumption of quercetin and other related substrates under anaerobic conditions in the presence of HNO-releasing compounds and the enzyme. As with dioxygenase activity, a nonenzymatic base-catalyzed reaction of quercetin with HNO isobserved above pH 7, but no enhancement of this basal reactivity is found upon addition of divalent metal salts. Unique and regioselective N-containing products are characterized by MS analysis for both the enzymatic and nonenzymatic reactions
Iron
Mn2+
Ni2+
Nickel
dioxygen shows two binding modes to the nickel ion, which can convert each other. Due to the overlap between the vacant d orbitals of nickel and the lone pair p orbitals of dioxygen and quercetin, electron transfer occurs from quercetin to dioxygen via the nickel center. Both dioxygen and quercetin can be activated by their binding to the nickel ion. The triplet reactant complex favors the catalytic reaction, and the whole reaction contains four elementary steps. A nonchemical process, the Op-Od bond rotation along the nickel center, is suggested to be rate-limiting with a free energy barrier of 19.9 kcal/mol
Zn2+
additional information
Co2+
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Co2+ salt addition increases the activity of quercetin 2,3-dioxygenase 24fold. The Escherichia coli cultures were grown at 37°C and 200 rpm for 6 h, induced with isopropyl beta-D-thiogalactopyanoside to a final concentraton of 50 mg/l in the presence of 10 microM CoCl2, and allow to grow additional 4 h at 25°C. The protein contains 0.65-0.8 atom of cobalt and 0.1 atom of iron per subunit.
Co2+
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supplementing the cultures of strain FLA with CoCl2 results in 1.6fold higher quercetinase activity in crude extracts
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probably belongs to the nonblue class, two atoms per molecule of enzyme
required, enzyme-bound, structure, overview. Manual docking, different geometries of the copper site
Cu2+
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required, mononuclear copper(II) active site, binding structure, X-ray diffraction and NMR analysis, overview. Direct coordinative interaction between copper(II) ion and the carboxylate group of Glu73. Complexes modeling, overview
Cu2+
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required, the copper ion is mainly coordinated by three His residues and a water molecule in a distorted tetrahedral geometry. In a minor form, the metal is penta-coordinated by three His, a glutamate, and an aquo ligand in a trigonal bipyramidal geometry. The major role of the activesite metal ion could be to correctly position the substrate and to stabilize transition states and intermediates rather than to mediate electron transfer
Cu2+
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required, a flavonolate ion (fla-, deprotonated substrate) is bound through the 3-hydroxy group to the copper(II) ion, which exhibits a distorted squarepyramidal geometry
Cu2+
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the enzyme has a mononuclear type 2 copper center, steric effects of the protein environment contribute to maintain the orientation of the substrate dissociated from the copper center. A prior rearrangement of the Cu2+-alkylperoxo complex and a subsequent hydrogen bond switching assisted by the movement of Glu73 can facilitate formation of an endoperoxide intermediate selectively
Cu2+
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Cu2+ salt addition increases the activity of quercetin 2,3-dioxygenase 1.4fold. The Escherichia coli cultures were grown at 37°C and 200 rpm for 6 h, induced with isopropyl beta-D-thiogalactopyanoside to a final concentraton of 50 mg/l in the presence of 10 microM CuCl2, and allow to grow additional 4 h at 25°C.
Cu2+
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activates, Cu-QDO, during the reaction mechanism of Cu-QDO dioxygen binds to the metal ion of the Cu-QDO-quercetin complex, yielding a Cu2+-superoxo quercetin radical intermediate, which then forms a Cu2+-alkylperoxo complex, the alkylperoxo complex evolves into endoperoxide intermediate that decomposes to the product
Cu2+
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required, the major role of the activesite metal ion could be to correctly position the substrate and to stabilize transition states and intermediates rather than to mediate electron transfer
different coordination geometry in the two active sites of the dimer
Iron
when the metal cofactor is replaced by an iron ion, the rate-limiting step switches from the Op-Od bond rotation to the collapse of the five-membered ring intermediate, corresponding to a free energy barrier of 30.3 kcal/mol
Mn2+
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Mn2+ salt addition increases the activity of quercetin 2,3-dioxygenase 35fold. The Escherichia coli cultures were grown at 37°C and 200 rpm for 6 h, induced with isopropyl beta-D-thiogalactopyanoside to a final concentraton of 50 mg/l in the presence of 10 microM MnSO4, and allow to grow additional 4 h at 25°C. The protein containes 1.6-1.9 atoms of Mn/subunit.
Mn2+
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activates, Mn-QDO, Mn2+ i the preferred metal ion. Mn-QDO in absence of O2 shows ability to react with nitroxyl (HNO)-singly reduced form of NO. HNO is incorporated into quercetin in the same manner as dioxygen, yet the reaction is strictly regioselective, as the only product is 2-((3,4-dihydroxyphenyl)(imino) methoxy)-4,6-dihydroxybenzoate
Ni2+
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Ni2+ salt addition increases the activity of quercetin 2,3-dioxygenase 2.6fold. The Escherichia coli cultures were grown at 37°C and 200 rpm for 6 h, induced with isopropyl beta-D-thiogalactopyanoside to a final concentraton of 50 mg/l in the presence of 10 microM NiCl2, and allow to grow additional 4 h at 25°C.
Ni2+
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supplementing the cultures of strain FLA with NiCl2 results in 6.1fold higher quercetinase activity in crude extracts
Ni2+
activates best, enzyme-bound, a nickel quercetinase. Ni2+ ions support correct folding, the catalytic activity of wild-type QueD is likely mediated by a Ni2+ center
additional information
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fungal quercetinases appear to exclusively utilize a Cu2+ ion for catalysis
additional information
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synthesis of a set of copper(II) complexes [CuIILn(AcO)] and their flavonolate adducts [CuIILn(fla)] with the series of carboxyl-group-containing ligands LnH, the treatment of the ligands with CuII(OAc)2xH2O gives the corresponding mononuclear copper(II) complexes [CuIILn(OAc)], ligand structures, mass spectrometric analysis, overview
additional information
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Cd2+ does not increase the activity of quercetin 2,3-dioxygenase. The Escherichia coli cultures were grown at 37°C and 200 rpm for 6 h, induced with isopropyl beta-D-thiogalactopyanoside to a final concentraton of 50 mg/l in the presence of 10 microM CdCl2, and allow to grow additional 4 h at 25°C.
additional information
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Fe2+ does not increase the activity of quercetin 2,3-dioxygenase. The Escherichia coli cultures were grown at 37°C and 200 rpm for 6 h, induced with isopropyl beta-D-thiogalactopyanoside to a final concentraton of 50 mg/l in the presence of 10 microM FeCl2, and allow to grow additional 4 h at 25°C.
additional information
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Zn2+ does not increase the activity of quercetin 2,3-dioxygenase. The Escherichia coli cultures were grown at 37°C and 200 rpm for 6 h, induced with isopropyl beta-D-thiogalactopyanoside to a final concentraton of 50 mg/l in the presence of 10 microM ZnSO4, and allow to grow additional 4 h at 25°C.
additional information
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the bacterial enzyme is capable of using different divalent metal ions for catalysis, with preference Mn2+, Co2+, Fe2+, Ni2+, Cu2+in descending order, suggesting that the redox properties of the metal are relatively unimportant for the catalytic reaction. The major role of the active site metal ion could be to correctly position the substrate and to stabilize transition states and intermediates rather than to mediate electron transfer. The recombinant enzyme is able to exchange its active-site metal ion while retaining catalytic activity
additional information
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the enzyme from Bacillus subtilis is active with several divalent metal cofactors such as Fe, Mn, and Co, although Mn(II) is the preferred cofactor for this enzyme
additional information
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QDO is a mononuclear metalloenzyme hosting various transition metal ions (Cu2+, Mn2+, Fe2+) in its active site depending on the origin of the protein, different metal complex structures, overview
additional information
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fungal quercetinases appear to exclusively utilize a Cu2+ ion for catalysis
additional information
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no increase in activity is observed when Mn2+, Fe2+, Cu2+, or Zn2+ is added to the culture medium
additional information
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the bacterial enzyme is capable of using different divalent metal ions for catalysis, with preference Ni2+, Co2+, Mn2+, Fe2+ in descending order, suggesting that the redox properties of the metal are relatively unimportant for the catalytic reaction. The major role of the active site metal ion could be to correctly position the substrate and to stabilize transition states and intermediates rather than to mediate electron transfer
additional information
the enzyme is metal-dependent. Cu2+ and Zn2+ do not support catalytic activity. Heterologous formation of catalytically active, native QueD holoenzyme requires Ni2+, Co2+ or Mn2+, i.e. metal ions that prefer an octahedral coordination geometry, and an intact 3His/1Glu motif or a 4His environment of the metal. The observed metal occupancies suggest that metal incorporation into QueD is governed by the relative stability of the resulting metal complexes, rather than by metal abundance. Ni2+ most likely is the physiologically relevant cofactor of QueD of Streptomyces sp. FLA, metal content analysis of wild-type and mutant enzymes, detailed overview