Information on EC 1.13.11.15 - 3,4-dihydroxyphenylacetate 2,3-dioxygenase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.13.11.15
-
RECOMMENDED NAME
GeneOntology No.
3,4-dihydroxyphenylacetate 2,3-dioxygenase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
3,4-dihydroxyphenylacetate + O2 = 2-hydroxy-5-carboxymethylmuconate semialdehyde
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oxidation
-
-
-
-
redox reaction
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-
-
-
reduction
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
4-hydroxyphenylacetate degradation
Tyrosine metabolism
-
-
Microbial metabolism in diverse environments
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-
SYSTEMATIC NAME
IUBMB Comments
3,4-dihydroxyphenylacetate:oxygen 2,3-oxidoreductase (decyclizing)
An iron protein.
CAS REGISTRY NUMBER
COMMENTARY hide
37256-56-7
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain 3B-1
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
Arthrobacter synephrinum
-
-
-
Manually annotated by BRENDA team
weak activity
-
-
Manually annotated by BRENDA team
C
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
strain CL-1, weak activity
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-
Manually annotated by BRENDA team
strain CL-1, weak activity
-
-
Manually annotated by BRENDA team
gene hpaD
-
-
Manually annotated by BRENDA team
CBS3
-
-
Manually annotated by BRENDA team
weak activity
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
-
In mutant Y257F, steps of the catalytic cycle are slowed by as much as 100fold by the mutation compared to the wild-type enzyme due to failure of mutant Y257F to facilitate the observed distortion of the bound 3,4-dihydroxyphenylacetate that is proposed to promote transfer of one electron to O2, Steady-state and transient kinetic analyses
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2,3-dihydroxybenzoate + O2
alpha-hydroxy-muconic semialdehyde
show the reaction diagram
-
-
mutant H200F produces alpha-carboxy-cis,cis-muconic acid
-
?
3,4 dihydroxyphenylacetate + O2
1-carboxymethyl-4-hydroxy-cis-muconic semialdehyde
show the reaction diagram
-
-
-
-
?
3,4-dihydroxybenzoic acid + O2
2-hydroxy-5-carboxymuconate semialdehyde
show the reaction diagram
3,4-dihydroxymandelate + O2
?
show the reaction diagram
3,4-dihydroxyphenylacetate + O2
1-carboxymethyl-4-hydroxy-cis-muconic semialdehyde
show the reaction diagram
-
-
-
-
?
3,4-dihydroxyphenylacetate + O2
2-hydroxy-5-carboxymethyl-cis,cis-muconate semialdehyde
show the reaction diagram
-
-
-
-
?
3,4-dihydroxyphenylacetate + O2
2-hydroxy-5-carboxymethylmuconate semialdehyde
show the reaction diagram
3,4-dihydroxyphenylacetate + O2
?
show the reaction diagram
3,4-dihydroxyphenylpropionate + O2
2-hydroxy-5-carboxyethylmuconate semialdehyde
show the reaction diagram
3-(3,4-dihydroxyphenyl)-DL-alanine + O2
(2E,4Z)-7-amino-5-formyl-2-hydroxyocta-2,4-dienedioic acid
show the reaction diagram
-
-
-
-
?
4-nitrocatechol + O2
?
show the reaction diagram
N-formyl-3-(3,4-dihydroxyphenyl)-DL-alanine + O2
(2E,4Z)-5-formyl-7-(formylamino)-2-hydroxyocta-2,4-dienedioic acid
show the reaction diagram
-
-
-
-
?
additional information
?
-
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
3,4 dihydroxyphenylacetate + O2
1-carboxymethyl-4-hydroxy-cis-muconic semialdehyde
show the reaction diagram
-
-
-
-
?
3,4-dihydroxyphenylacetate + O2
2-hydroxy-5-carboxymethylmuconate semialdehyde
show the reaction diagram
additional information
?
-
-
LB400 cells grown with 3-hydroxyphenylacetate degrade homoprotocatechuate and show homoprotocatechuate 2,3-dioxygenase activity
-
-
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Fe
-
contains an active site Fe(II)
Fe3+
-
nonphysiological metal, incorporated in Fe-MndD
Manganese
Mg2+
-
1.1-1.3 gatom of Mg2+ per mol of enzyme, essential for activity
Mn
-
wild-type enzyme amd mutant enzymes H200A and H200Q: 0.6 Mn per monomer. Mutant enzyme H200E: 0.3 Mn per monomer. Mutent enzyme H200N: 0.4 MN per monomer
NO
-
NO binding to a non-heme enzyme containing manganese allows examination of the factors governing the formation and detection of the MIII-O2.- species in all forms of th enzyme. NO, and presumably O2, binding is sensitive to both the nature of the catecholic substrate present and the nature of the active-site amino acid residue at position 200, spectral analysis, overview
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,10-phenanthroline
3,4-dichlorophenylacetate
-
5 mM, 50% inhibition
3,4-dihydroxybenzoate
3,4-dihydroxycinnamate
-
-
3,4-Dihydroxyphenylpropionate
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-
4-Methylcatechol
-
-
4-nitrocatechol
5,5'-dithiobis-(2-nitrobenzoate)
Arthrobacter synephrinum
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1 mM, complete inhibition
8-hydroxyquinoline
air
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initially high activity of Fe-MndD, becomes inactivtated within one h
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Copper salicylate
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0.166 mM, rate of oxygen uptake is decreased by 38%, rate of product formation is diminished by 58%
Cu2+
-
0.01 mM, 80% inhibition
dithiothreitol
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0.1 mM, 13% inhibition
ferricyanide
Ferrous ammonium sulfate
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0.1 mM, 32% inhibition
K3Fe(CN)6
-
10 mM, 35% inhibition
m-hydroxyphenylacetate
-
-
p-hydroxymercuribenzoate
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0.1 mM, 13% inhibition
p-hydroxyphenylacetate
-
-
succinate semialdehyde
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5 mM 65% inhibition
Zn2+
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0.01 mM, complete inhibition
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Sodium ascorbate
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1 mM, revers purification inactivation of Fe-MndD, about 30-35% higher catalytic activity than the purified enzyme
additional information
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.074 - 0.45
3,4-dihydroxybenzoic acid
0.5
3,4-dihydroxymandelate
Arthrobacter synephrinum
-
-
0.0004 - 0.035
3,4-Dihydroxyphenylacetate
0.0037
3,4-Dihydroxyphenylpropionic acid
-
32C
0.015
4-nitrocatechol
-
pH 7.5, 22C
0.035
DL-3,4-dihydroxymandelic acid
-
32C
0.037 - 0.062
O2
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.01 - 0.02
2,3-Dihydroxybenzoate
0.067 - 10
3,4-Dihydroxyphenylacetate
0.05 - 0.4
4-nitrocatechol
additional information
additional information
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.3
3,4-dihydroxybenzoate
-
-
0.3
3,4-dihydroxycinnamate
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-
1
3,4-Dihydroxyphenylpropionate
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-
1.5
4-Methylcatechol
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-
0.5
4-nitrocatechol
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-
3.8
catechol
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-
0.27
m-hydroxyphenylacetate
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-
1.8
p-hydroxyphenylacetate
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-
additional information
additional information
-
Ki-values for 4-nitrocatechol for the H200A, H200E, H200N and H200Q variants are between 0.015 and 0.025 mM
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2.51
Arthrobacter synephrinum
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-
6.2
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recombinant enzyme
additional information
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-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.5 - 7.5
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assay at
6.9
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substrate 4-nitrocatechol
7 - 8
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mutant enzyme H200N and H200Q
7.2
Arthrobacter synephrinum
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7.2 - 7.8
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8 - 9
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wild-type enzyme
8.4
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substrate 3,4-dihydroxyphenylacetate
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.1 - 9.3
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78% of maximal activity at pH 7.1 and 9.3
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 22
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assay at 4C or 22C
22
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assay at room temperature
50
Arthrobacter synephrinum
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-
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
40 - 64
-
40C: about 40% of maximal activity, 64C: about 75% of maximal activity
PDB
SCOP
CATH
ORGANISM
UNIPROT
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
33000
-
gel filtration
102000
-
gel filtration
106000
-
gel filtration
120000
-
equilibrium sedimentation
138000
-
gel filtration
140000
-
gel filtration
220000
-
equilibrium sedimentation
282000
Arthrobacter synephrinum
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hexamer
-
6 * 42500, or 42500, SDS-PAGE
homotetramer
tetramer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystallizations of wild-type FeHPCD and mutant Y257F with bound substrates 3,4-dihydroxyphenylacetate and 4-nitrocatechol, hanging drop vapour diffusion method, in either 13% PEG 6000, 0.1 M calcium chloride, 0.1 M Tris-HCl, pH 6.5-7.4 or in 18% PEG 8000, 0.1 M calcium acetate, 0.1 M sodium cacodylate, pH 6.5, at 20C, cryoprotectant solution containing either 20-25% PEG400 or 20% glycerol in the mother liquor solution, X-ray diffraction structure determination and analysis at 1.50-1.60 A resolution
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
-
most stable at
439614
7.5
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maximal stability at
439626
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0
-
pH 7.5, 0.1 M sodium potassium phosphate, 1 mM DTT, complete loss of activity after 48 h
40
-
half-life: 13 min
50
Arthrobacter synephrinum
-
30 min, stable
55
-
5 min, stable
63
-
2 h, 5% loss of activity
88
-
10 min, 50% loss of activity
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
dialysis against buffer for several hours at 0C, no appreciable loss of activity
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Fe-HPCD stable in air for at least several days
-
inactivation by 6 M guanidine hydrochloride
Arthrobacter synephrinum
-
stabilization in presence of low concentrations of acetone or ethanol
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
10C, lyophilized preparations of the enzyme, 8 months, stable
-
room temperature, little loss of activity after 3 months
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
partial
recombinant enzyme from Escherichia coli
-
recombinant wild-type and mutant Y257F enzymes by a procedure including anion exchange chromatography
-
wild-type and mutant enzymes
-
wild-type enzyme and H200 mutants
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli BL21
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expression in Escherichia coli DH5alpha
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gene hpaD, gene cluster hpaG1G2EDFHI encodes the homoprotocatechuate pathway is located at C2 in strain LB400 genome, quantitative expression analysis
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overexpression in Escherichia coli
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recombinant expression of wild-type and mutant Y257F enzymes
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wild-type and mutant enzymes expressed in Escherichia coli
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
induction of fumarylacetoacetate hydrolase HmgB in Burkholderia xenovorans LB400 during growth on hydroxyphenylacetates
-
LB400 cells grown with 3-hydroxyphenylacetate and 4-hydroxyphenylacetate both show induction of the homoprotocatechuate pathway, they degrade homoprotocatechuate and show homoprotocatechuate 2,3-dioxygenase activity. The hmgA1 gene is upregulated by 3-hydroxyphenylacetate in strain LB400. The expression of the hpaD gene, encoding HPC 2,3-dioxygenase, is induced in 3-hydroxyphenylacetate and 4-hydroxyphenylacetate grown-cells
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E266Q
-
specific activity is less than 0.1% of that of the wild-type enzyme, binds 33% of the wild-type level of manganese
H155A
-
specific activity is less than 0.1% of that of the wild-type enzyme, binds 0.4% of the wild-type level of manganese
H200A
-
KM-value for 3,4-dihydroxyphenylacetate is 1.75fold lower than wild-type value, kcat for 3,4-dihydroxyphenylacetate is 69fold lower than wild-type value
H200E
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KM-value for 3,4-dihydroxyphenylacetate is 1.2fold lower than wild-type value, kcat for 3,4-dihydroxyphenylacetate is more than 276fold lower than wild-type value, Mn content per monomer is 2fold lower than in wild-type enzyme
H200N
-
KM-value for 3,4-dihydroxyphenylacetate is identical to wild-type value, kcat for 3,4-dihydroxyphenylacetate is 1.73fold lower than wild-type value, Mn content per monomer is 65% of wild-type value
H200Q
-
KM-value for 3,4-dihydroxyphenylacetate is 1.2fold lower than wild-type value, kcat for 3,4-dihydroxyphenylacetate is 1.3fold lower than wild-type value
H214A
-
specific activity is less than 0.1% of that of the wild-type enzyme, binds 1.8% of the wild-type level of manganese
H42A
-
97% of wild-type activity at 18C, 30% of wild-type activity at 37C
E266Q
-
specific activity is less than 0.1% of that of the wild-type enzyme, binds 33% of the wild-type level of manganese
-
H155A
-
specific activity is less than 0.1% of that of the wild-type enzyme, binds 0.4% of the wild-type level of manganese
-
H200A
-
KM-value for 3,4-dihydroxyphenylacetate is 1.75fold lower than wild-type value, kcat for 3,4-dihydroxyphenylacetate is 69fold lower than wild-type value
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H200E
-
KM-value for 3,4-dihydroxyphenylacetate is 1.2fold lower than wild-type value, kcat for 3,4-dihydroxyphenylacetate is more than 276fold lower than wild-type value, Mn content per monomer is 2fold lower than in wild-type enzyme
-
H200N
-
KM-value for 3,4-dihydroxyphenylacetate is identical to wild-type value, kcat for 3,4-dihydroxyphenylacetate is 1.73fold lower than wild-type value, Mn content per monomer is 65% of wild-type value
-
H200Q
-
KM-value for 3,4-dihydroxyphenylacetate is 1.2fold lower than wild-type value, kcat for 3,4-dihydroxyphenylacetate is 1.3fold lower than wild-type value
-
H214A
-
specific activity is less than 0.1% of that of the wild-type enzyme, binds 1.8% of the wild-type level of manganese
-
H42A
-
97% of wild-type activity at 18C, 30% of wild-type activity at 37C
-
H200A
-
Km-value for 3,4-dihydroxyphenylacetate is 6fold lower than wild-type value, kcat for 3,4-dihydroxyphenylacetate is 33.3fold lower than wild-type value
H200E
-
Km-value for 3,4-dihydroxyphenylacetate is 75fold lower than wild-type value, kcat for 3,4-dihydroxyphenylacetate is 11.1fold lower than wild-type value
H200Q
-
Km-value for 3,4-dihydroxyphenylacetate is similar to wild-type value, kcat for 3,4-dihydroxyphenylacetate is 2.5fold lower than wild-type value
additional information
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