Information on EC 1.13.11.66 - hydroquinone 1,2-dioxygenase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.13.11.66
-
RECOMMENDED NAME
GeneOntology No.
hydroquinone 1,2-dioxygenase
-
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
benzene-1,4-diol + O2 = (2Z,4E)-4-hydroxy-6-oxohexa-2,4-dienoate
show the reaction diagram
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
4-hydroxyacetophenone degradation
-
-
4-nitrophenol degradation I
-
-
Chlorocyclohexane and chlorobenzene degradation
-
-
Microbial metabolism in diverse environments
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-
SYSTEMATIC NAME
IUBMB Comments
benzene-1,4-diol:oxygen 1,2-oxidoreductase (decyclizing)
The enzyme is an extradiol-type dioxygenase, and is a member of the nonheme-iron(II)-dependent dioxygenase family. It catalyses the ring cleavage of a wide range of hydroquinone substrates to produce the corresponding 4-hydroxymuconic semialdehydes.
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
C1I210; C1I209;
the two-subunit hydroquinone 1,2-dioxygenase PnpCD is the ring cleavage enzyme in para-nitrophenol catabolism
physiological function
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-difluorohydroquinone + O2
?
show the reaction diagram
2,5-difluorohydroquinone + O2
?
show the reaction diagram
2,5-dimethylhydroquinone + O2
2-methylmaleylacetone
show the reaction diagram
2,6-dibromohydroquinone + O2
2-bromomaleylacetate + Br-
show the reaction diagram
-
-
-
?
2,6-dichloro-p-hydroquinone + O2
2-chloromaleylacetate + Cl-
show the reaction diagram
-
-
-
?
2,6-dichlorohydroquinone + O2
2-chloromaleylacetate + Cl-
show the reaction diagram
complete conversion of substrate
-
-
?
2,6-dichlorohydroquinone + O2
?
show the reaction diagram
-
-
0.5-0.6 equiv. of chloride is released during turnover of substrate
-
?
2,6-dimethylhydroquinone + O2
2-methylmaleylacetone + ?
show the reaction diagram
-
-
-
?
2-(1-methyl1-octyl)-hydroquinone + O2
?
show the reaction diagram
2-chloro-6-methylhydroquinone
?
show the reaction diagram
-
complete conversion of substrate, yields a mixture of 1,2- and 1,6-cleavage products. The two modes of cleavage have different Km values for oxygen, consistent with a mechanism in which the substrate binds in two catalytically productive orientations
-
?
2-chlorohydroquinone + O2
?
show the reaction diagram
2-ethylhydroquinone + O2
?
show the reaction diagram
2-hexylhydroquinone + O2
?
show the reaction diagram
2-methoxyhydroquinone + O2
?
show the reaction diagram
59% of the activity with hydroquinone
-
-
?
2-methylhydroquinone + O2
?
show the reaction diagram
2-methylhydroquinone + O2
maleylacetone
show the reaction diagram
-
-
-
-
?
2-pentylhydroquinone + O2
?
show the reaction diagram
19% of the activity with hydroquinone
-
-
?
2-propylhydroquinone + O2
?
show the reaction diagram
23% of the activity with hydroquinone
-
-
?
2-tert-butylhydroquinone + O2
?
show the reaction diagram
5% of the activity with hydroquinone
-
-
?
3,5-difluorohydroquinone + O2
?
show the reaction diagram
4-nitrophenol + O2
?
show the reaction diagram
C1I210; C1I209;
-
-
-
?
benzene-1,4-diol + O2
(2Z,4E)-4-hydroxy-6-oxohexa-2,4-dienoate
show the reaction diagram
bromohydroquinone + O2
?
show the reaction diagram
bromohydroquinone + O2
maleylacetate + Br-
show the reaction diagram
-
-
-
?
chlorohydroquinone + O2
?
show the reaction diagram
-
70% of the activity with hydroquinone
-
-
?
chlorohydroquinone + O2
maleylacetate + Cl-
show the reaction diagram
-
-
-
?
hydroquinone + O2
4-hydroxymuconic acid semialdehyde
show the reaction diagram
hydroquinone + O2
4-hydroxymuconic semialdehyde
show the reaction diagram
hydroquinone + O2
gamma-hydroxymuconic acid semialdehyde
show the reaction diagram
methoxyhydroquinone + O2
?
show the reaction diagram
-
50% of the activity with hydroquinone
-
-
?
methylhydroquinone + O2
?
show the reaction diagram
methylhydroquinone + O2
maleylacetone + ?
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
2-methylhydroquinone + O2
?
show the reaction diagram
4-nitrophenol + O2
?
show the reaction diagram
C1I210 AND C1I209
-
-
-
?
benzene-1,4-diol + O2
(2Z,4E)-4-hydroxy-6-oxohexa-2,4-dienoate
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
copper
-
copper content is 0.12 atoms per monomer
Fe3+
C1I210; C1I209;
binding structure, overview
Manganese
-
preincubation in presence of 0.1 mM Fe2+ and 1 mM Mn2+ increases activity by 10fold
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2,2'-dipyridyl
-
inactivation
2,6-dibromophenol
-
competitive
3,4-dihydroxybenzoate
0.2 mM, 94% inhibition; 0.2 mM, 94% inhibition
3-bromocatechol
-
inactivation
4-coumaric acid
0.2 mM, 97% inhibition; 0.2 mM, 97% inhibition
4-hydroxybenzoate
4-Hydroxybenzonitrile
-
0.2 mM, no residual activity
4-hydroxycinnamate
-
0.2 mM, no residual activity
4-nitrophenol
bromohydroquinone
-
substrate inhibition
caffeic acid
0.2 mM, 98% inhibition; 0.2 mM, 98% inhibition
catechol
0.2 mM, 93% inhibition; 0.2 mM, 93% inhibition
chlorohydroquinone
-
substrate inhibition
hydrogen peroxide
-
inactivation
Hydroxyhydroquinone
-
0.2 mM, 9% residual activity
methoxyhydroquinone
-
strong substrtae inhibition
o-phenanthroline
-
inactivation
ortho-disubstituted phenols
-
-
-
phenol
0.2 mM, 98% inhibition; 0.2 mM, 98% inhibition
resorcinol
0.2 mM, 99% inhibition; 0.2 mM, 99% inhibition
vanillate
0.2 mM, 86% inhibition; 0.2 mM, 86% inhibition
Vanillyl alcohol
0.2 mM, 62% inhibition; 0.2 mM, 62% inhibition
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
-
ligandation with 4-hydroxybenzoate prevents the enzyme from irreversible inactivation
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0013
2,6-dibromohydroquinone
-
pH 7.0, 23°C
0.0135
2,6-dichloro-p-hydroquinone
pH 6.7, 22°C
0.0032
2,6-dichlorohydroquinone
-
pH 7.0, 23°C
0.055
2,6-dimethylhydroquinone
-
pH 7.0, 23°C
0.0044
2-chloro-6-methylhydroquinone
-
pH 7.0, 23°C
0.2
2-methylhydroquinone
-
pH 6.4, 30°C, recombinant enzyme
-
0.0085
bromohydroquinone
-
pH 7.0, 23°C
0.01
chlorohydroquinone
-
pH 7.0, 23°C
0.0022 - 0.255
hydroquinone
0.041
methylhydroquinone
-
pH 7.0, 23°C
0.021 - 0.26
O2
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.61
2,6-dibromohydroquinone
-
pH 7.0, 23°C
2.1
2,6-dichloro-p-hydroquinone
pH 6.7, 22°C
3.05
2,6-dichlorohydroquinone
-
pH 7.0, 23°C
1.25
2,6-dimethylhydroquinone
-
pH 7.0, 23°C
2.41
2-chloro-6-methylhydroquinone
-
pH 7.0, 23°C
2.1 - 13.5
hydroquinone
1.4 - 4.8
O2
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1200
2,6-dibromohydroquinone
-
pH 7.0, 23°C
1000
2,6-dichlorohydroquinone
-
pH 7.0, 23°C
23
2,6-dimethylhydroquinone
-
pH 7.0, 23°C
540
2-chloro-6-methylhydroquinone
-
pH 7.0, 23°C
6136
hydroquinone
pH 7.0, 25°C; pH 7.0, 25°C
0.7 - 26
O2
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.014
4-hydroxybenzoate
-
pH 7.0, 25°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.27
crude extract, substrate hydroquinone, pH 7.0, 37°C
0.88
crude extract, substrate 2-chlorohydroquinone, pH 7.0, 37°C
3.1
-
pH 7.4, temperature not specified in the publication
5.89
-
pH 7.0, 25°C
6.1
pH 7.0, 25°C; pH 7.0, 25°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.4
-
with substrate 2-methylhydroquinone
6.7
optimal in 20 mM potassium phosphate buffer
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3 - 10
-
-
7
-
20% of maximum activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30
-
assay at
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20 - 70
-
-
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
17800
-
2 * 17800, alpha-subunit, + 2 * 38300, beta-subunit
18000
-
SDS-PAGE
34000
-
x * 34000, SDS-PAGE
36000
x * 36000, SDS-PAGE
36522
-
x * 36522, electrospray LC-MS
38300
-
2 * 17800, alpha-subunit, + 2 * 38300, beta-subunit
112000
-
gel filtration
120000
gel filtration; gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
heterotetramer
tetramer
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified full-length or proteolytically truncated PnpCD in apo form and in complex with Fe3+ or substrate analogue hydroxybenzonitrile and Cd2+, i.e. apo-PnpCD, PnpCD-Fe3+, and PnpCD-Cd2+-HBN, sitting-drop vapor diffusion method, 15-20 mg/ml protein in 10mM Tris-HCl, pH 8.0, and 100 mM NaCl, is mixed with reservoir solution containing 0.2 M sodium thiocyanate, 20% w/v PEG 3350, 20°C, method optimization, X-ray diffraction structure determination and analysis
C1I210; C1I209;
generation of a homology model, based on zinc protein PDB entry 1ZSW, which predicts that the tertiary structure of the enzyme differs significantly from that of the extradiol dioxygenases, and that the residues ligating the Fe(II) are H11, H227, and E276
-
purified enzyme free or in complex with substrate methylhydroquinone under anaerobic conditions, or with inhibitors 4-hydroxybenzoate and 4-nitrophenol, sitting drop vapor diffusion method, mixing of 0.001 ml of 8 mg/ml protein in 25 mM Tris, pH 7.0, 5 mM NaCl, and 0.5 mM ligand, with 0.001 ml of reservoir solution containing 14% PEG 3350, 0.35 M MgCl2, and 0.1 M MES, pH 6.5, 4°C, 1 day, the structure of the free enzyme is obtained by soaking the crystals in a stabilizing solution containing 16% PEG 3350, 0.35 M MgCl2, and 0.1 M MES, pH 6.5, for two days, changing the solution three times, in order to remove the ligands, X-ray diffraction structure determination and analysis at 1.90-2.40 resolution, molecular replacement using the coordinates of PnpCD structure from Pseudomonas sp. strain WBC-3, PDB ID 4ZXA as template, modeling
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3
-
20% activity after 30 min at pH 3.0
719179
10
-
60% activity after 30 min at pH 10.0
719179
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60
-
the purified enzyme retains 35% activity after 20 min at 60°C
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
4-hydrobenzoate, the competitive inhibitor of the heterotetrameric HapCD hydroquinone dioxygenase of Pseudomonas fluorescens ACB, stabilizes the enzyme
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20°C, 50% glycerol, 48% residual activity after 30 days
-
-70°C, 50% glycerol, 77% residual activity after 30 days
-
fairly stable for several hours on ice and retains approximately 80% of activity after 72 h
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ligandation with 4-hydroxybenzoate prevents the enzyme from irreversible inactivation
-
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration
C1I210; C1I209;
recombinant N-terminally His6-tagged enzyme from Escherichia coli by nickel affinity chromatography, tag removal by rTEV protease cleavage, over 95% purity. No enzyme activity can be detected with purified YaiA, even when it is purified in the presence of 4-hydrobenzoate
-
using Ni-NTA chromatography
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli as His-tagged fusion proteins
-
expression in Escherichia coli
gene yaiA, encoded in the the yahCD-yaiAB operon, DNA and amino acid sequence determination and analysis, recombinant expression of N-terminally His6-tagged enzyme in Escherichia coli. YaiA activity could be detected in crude Escherichia coli cytoplasmic extract overexpressing Lactcoccus lactis YaiA from a plasmid
-
genes pnpCD, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3)
C1I210; C1I209;
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
the the yahCD-yaiAB operon is induced by copper but not by quinones
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E248Q
C1I210; C1I209;
site-directed mutagenesis, the mutation results in complete loss of enzyme activity
F264A
C1I210; C1I209;
site-directed mutagenesis, the mutation results in almost complete loss of enzyme activity
F79A
C1I210; C1I209;
site-directed mutagenesis, the mutation results in almost complete loss of enzyme activity
L252A
C1I210; C1I209;
site-directed mutagenesis, the mutation results in a 70% loss of enzyme activity
L313A
C1I210; C1I209;
site-directed mutagenesis, the mutation results in almost complete loss of enzyme activity
V315A
C1I210; C1I209;
site-directed mutagenesis, the mutation results in a 50% loss of enzyme activity
W230A
C1I210; C1I209;
site-directed mutagenesis, the mutation results in a 70% loss of enzyme activity
W273a
C1I210; C1I209;
site-directed mutagenesis, the mutation results in almost complete loss of enzyme activity
W76A
C1I210; C1I209;
site-directed mutagenesis, the mutation results in almost complete loss of enzyme activity
E276A
-
less than 6% of wild-type activity
H11A
-
less than 6% of wild-type activity
H159A
-
67% of wild-type activity
H227A
-
less than 6% of wild-type activity
Y266F
-
about 6% of wild-type activity
E278A
mutation in putative Fe(II)-binding site, complete loss of activity
H162A
mutation in putative Fe(II)-binding site, complete loss of activity
H229A
mutation in putative Fe(II)-binding site, complete loss of activity
additional information