Information on EC 1.23.5.1 - violaxanthin de-epoxidase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.23.5.1
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RECOMMENDED NAME
GeneOntology No.
violaxanthin de-epoxidase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
antheraxanthin + L-ascorbate = zeaxanthin + L-dehydroascorbate + H2O
show the reaction diagram
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-
-
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violaxanthin + 2 L-ascorbate = zeaxanthin + 2 L-dehydroascorbate + 2 H2O
show the reaction diagram
overall reaction
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-
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violaxanthin + L-ascorbate = antheraxanthin + L-dehydroascorbate + H2O
show the reaction diagram
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
carotenoid biosynthesis
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Carotenoid biosynthesis
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Metabolic pathways
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Biosynthesis of secondary metabolites
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SYSTEMATIC NAME
IUBMB Comments
violaxanthin:ascorbate oxidoreductase
Along with EC 1.14.13.90, zeaxanthin epoxidase, this enzyme forms part of the xanthophyll (or violaxanthin) cycle for controlling the concentration of zeaxanthin in chloroplasts. It is activated by a low pH of the thylakoid lumen (produced by high light intensity). Zeaxanthin induces the dissipation of excitation energy in the chlorophyll of the light-harvesting protein complex of photosystem II. In higher plants the enzyme reacts with all-trans-diepoxides, such as violaxanthin, and all-trans-monoepoxides, but in the alga Mantoniella squamata, only the diepoxides are good substrates.
CAS REGISTRY NUMBER
COMMENTARY hide
57534-73-3
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
var. Romaine
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Manually annotated by BRENDA team
cultivar Jyothi
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Manually annotated by BRENDA team
L. cv. Kleine Rheinländerin
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
malfunction
a reduction in enzyme expression results in greater photosensitivity
metabolism
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the xanthophyll cycle is an important photoprotective process functioning in plants. One of its forms, the violaxanthin cycle, involves interconversion between violaxanthin, antheraxanthin, and zeaxanthin
physiological function
additional information
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molecular dynamics and simulations, overview
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
all-trans-neoxanthin + ascorbate
? + dehydroascorbate + H2O
show the reaction diagram
antheraxanthin + acceptor
zeaxanthin + reduced acceptor
show the reaction diagram
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-
-
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?
antheraxanthin + ascorbate
zeaxanthin + dehydroascorbate + H2O
show the reaction diagram
antheraxanthin + L-ascorbate
zeaxanthin + L-dehydroascorbate + H2O
show the reaction diagram
cryptoxanthin epoxide + ascorbate
?
show the reaction diagram
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92% of the activity with violaxanthin
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?
cryptoxanthin-5,6,5',6'-di-epoxide + ascorbate
? + dehydroascorbate + H2O
show the reaction diagram
cryptoxanthin-5,6-epoxide + ascorbate
? + dehydroascorbate + H2O
show the reaction diagram
diadinoxanthin + ascorbate
?
show the reaction diagram
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the activity is 2.25fold higher than the activity of violaxanthin
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?
diadinoxanthin + ascorbate
? + dehydroascorbate + H2O
show the reaction diagram
lutein epoxide + ascorbate
?
show the reaction diagram
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the activity is 1.33fold higher than the activity of violaxanthin
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-
?
lutein-5,6-epoxide + ascorbate
? + dehydroascorbate + H2O
show the reaction diagram
neoxanthin + ascorbate
?
show the reaction diagram
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10% of the activity with violaxanthin
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-
?
violaxanthin + 2 L-ascorbate
zeaxanthin + 2 L-dehydroascorbate + 2 H2O
show the reaction diagram
violaxanthin + acceptor
antheraxanthin + reduced acceptor
show the reaction diagram
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-
-
-
?
violaxanthin + ascorbate
antheraxanthin + dehydroascorbate + H2O
show the reaction diagram
violaxanthin + L-ascorbate
antheraxanthin + L-dehydroascorbate + H2O
show the reaction diagram
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
antheraxanthin + ascorbate
zeaxanthin + dehydroascorbate + H2O
show the reaction diagram
antheraxanthin + L-ascorbate
zeaxanthin + L-dehydroascorbate + H2O
show the reaction diagram
violaxanthin + 2 L-ascorbate
zeaxanthin + 2 L-dehydroascorbate + 2 H2O
show the reaction diagram
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-
-
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?
violaxanthin + ascorbate
antheraxanthin + dehydroascorbate + H2O
show the reaction diagram
violaxanthin + L-ascorbate
antheraxanthin + L-dehydroascorbate + H2O
show the reaction diagram
additional information
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Cys
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2.7 mM, 50% inhibition
dithiothreitol
mercaptoethanol
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0.68 mM, 50% inhibition
o-phenanthroline
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0.025 mM, 50% inhibition. The rate of conversion of violaxanthin to antheraxanthin is relatively unchanged whereas the conversion of antheraxanthin to zeaxanthin is 50-80% inhibited
Pepstatin
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50% inhibition at 0.12 mM, reversible, protonation-induced structural change of the enzyme
pepstatin A
zeaxanthin
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product inhibition
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
digalactosyldiacylglyceride
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0.0067 mM, supports slow but nevertheless complete to nearly complete de-epoxidation
Lipid
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the enzyme requires lipid inverted hexagonal structures for activity
monogalactosyldiacylglyceride
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0.01 mM, supports rapid and complete de-epoxidation
monogalactosyldiacylglycerol
phosphatidylcholine
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supports slow and incomplete de-epoxidation
additional information
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0016 - 5.3
antheraxanthin
2.3 - 4.4
ascorbate
0.000049 - 11.1
violaxanthin
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.015
pH 7.4, 25°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 5.2
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irrespective of the presence of high or low ascorbate concentrations
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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dicyclohexylcarbodiimide alters the pH dependence of violaxanthin de-epoxidation
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.95
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isoelectric focusing
5.4
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isoelectric focusing
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
no protein detectable in root
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
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no expression in the cytoplasm or cell membrane of cucumber cotyledon protoplasts
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Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
45800
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gel filtration
60000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
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1 * 43300, SDS-PAGE
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
proteolytic modification
deduced amino acid sequence consists of a mature protein and a transit peptide of 103 amino acids
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5
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no activity above pH 6.5
676343
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
in presence of Tween 20. The enzyme has more than one disulfide bond and takes multiple forms depending on the extent of the reduction
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native enzyme partially by preparation of thylakoid membranes, further purifications of the VDE by, i.e. anion exchange chromatography and gel filtration, lead to a significant loss of VDE activity
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nickel affinity column chromatography
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TALON metal affinity resin column chromatography
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
a full-length violaxanthin de-epoxidase and deletion mutants of the N- and C-terminal regions are expressed in Escherichia coli and Nicotiana tabacum L. cv. Xanthi. High expression of the enzyme in Escherichia coli is achieved after adding the argU gene that encodes the Escherichia coli arginine AGA tRNA. The specific activity of the violaxanthin de-epoxidase expressed in Escherichia coli is low, possibly due to incorrect folding. The transformed tobacco exhibits a 13fold to 19fold increase in VDE specific activity, indicating correct protein folding
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DNA and amino acid sequence determination and analysis, phylogenetic analysis, transiently expression of GFP-tagged enzyme in cucumber protoplasts, anti-sense expression of CsVDE in Arabidopsis thaliana leading to silencing of the single copy Arabidopsis thaliana gene, quantitative real-time PCR expression analysis
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expressed in Escherichia coli Origami B cells
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expressed in Lycopersicom esculentum
expressed in Nicotiana tabacum
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expressed in Nicotiana tabacum NC89
expression in Escherichia coli
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
chilling conditions reduce the activity of the enzyme
CsVDE is quickly induced by cold and drought stress
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enzyme activation is triggered by a pH reduction in the thylakoids lumen occurring under saturating light
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expression is pregulated under high light conditions
VDE activity increases in greenhouse (0.15-2 mM/m2/s), shade (0.6-0.8 mN/m2/s) and sun (1.2-2.2 mM/m2/s) grown plants after one day of sun exposure as compared to prior to transfer to direct sunlight
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D114A
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inactive
D114N
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inactive
D117A
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site-directed mutagenesis, the mutant shows 60% reduced activity compared to the wild-type enzyme; the mutant shows 40% of wild type activity
D177A
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inactive
D177N
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inactive
D178A
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the mutant shows 56% activity compared to the wild type enzyme
D206I
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site-directed mutagenesis, the mutant shows 56% reduced activity compared to the wild-type enzyme; the mutant shows 44% of wild type activity
D86A
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site-directed mutagenesis, the mutant shows 19% reduced activity compared to the wild-type enzyme; the mutant shows 86% of wild type activity
D98L
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site-directed mutagenesis, the mutant shows 41% reduced activity compared to the wild-type enzyme; the mutant shows 59% of wild type activity
D98L/D117A
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site-directed mutagenesis, the mutant shows 59% reduced activity compared to the wild-type enzyme; the mutant shows 41% of wild type activity
D98L/D117A/D206I
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site-directed mutagenesis, the mutant shows 84% reduced activity compared to the wild-type enzyme; the mutant shows 16% of wild type activity
D98L/D117A/D206I/H168A
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site-directed mutagenesis, the mutant shows 94% reduced activity compared to the wild-type enzyme; the mutant shows 6% of wild type activity
DELTA1-4
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removal of 4 amino acids from the N-terminal region abolishes all violaxanthin de-epoxidase activity
DELTA258-349
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71 C-terminal amino acid can be removed without affecting activity
F123A
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the mutant shows 34% activity compared to the wild type enzyme
F155A
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the mutant shows 5% activity compared to the wild type enzyme
H121A
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the mutant shows 5% activity compared to the wild type enzyme
H168A
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site-directed mutagenesis, the mutant shows 80% reduced activity compared to the wild-type enzyme; the mutant shows 20% of wild type activity
N167A
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the mutant shows 121% activity compared to the wild type enzyme
Q119A
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the mutant shows 32% activity compared to the wild type enzyme
Q153A
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the mutant shows 75% activity compared to the wild type enzyme
Q153E
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the mutant shows 60% activity compared to the wild type enzyme
Q153L
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the mutant shows 30% activity compared to the wild type enzyme
T245A
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the mutant shows 82% activity compared to the wild type enzyme
W179A
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the mutant shows less than 2% activity compared to the wild type enzyme
W179N
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the mutant shows less than 2% activity compared to the wild type enzyme
Y175F
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the mutant shows 62% activity compared to the wild type enzyme
Y198F
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inactive
Y214F
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the mutant shows less than 2% activity compared to the wild type enzyme
H121A/H124A
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considerably lower pH dependence for binding than wild-type, cooperativity value around 2 compared to wild-type value of 3.7. Km-value for ascorbate is 3.2 mM compared to 1.9 mM for wild-type enzyme
H121R/H124R
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inactive mutant enzyme
H124R
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considerably lower pH dependence for binding than wild-type, cooperativity value around 2 compared to wild-type value of 3.7.Km-value for ascorbate is 1.5 mM compared to 1.9 mM for wild-type enzyme; considerably lower pH dependence for binding than wild-type, cooperativity value around 2 compared to wild-type value of 3.7. Km-value for ascorbate is 2.1 mM compared to 1.9 mM for wild-type enzyme
H134A
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considerably lower pH dependence for binding than wild-type, cooperativity value around 2 compared to wild-type value of 3.7
H167A/H173A
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considerably lower pH dependence for binding than wild-type, cooperativity value of 1.6 compared to wild-type value of 3.7. Km-value for ascorbate is 8.3 mM compared to 1.9 mM for wild-type enzyme
H167R/H173R
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considerably lower pH dependence for binding than wild-type, cooperativity value around 2 compared to wild-type value of 3.7. Km-value for ascorbate is 6.3 mM compared to 1.9 mM for wild-type enzyme
additional information
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enzyme downregulation by anti-sense expression of CsVDE in Arabidopsis thaliana showing reduced enzyme activity under high light stress