Information on EC 1.11.1.16 - versatile peroxidase

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria

EC NUMBER
COMMENTARY hide
1.11.1.16
-
RECOMMENDED NAME
GeneOntology No.
versatile peroxidase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
1-(4-hydroxy-3-methoxyphenyl)-2-(2-methoxyphenoxy)propane-1,3-diol + H2O2 = 4-hydroxy-3-methoxybenzaldehyde + 2-methoxyphenol + glycolaldehyde + H2O
show the reaction diagram
2 manganese(II) + 2 H+ + H2O2 = 2 manganese(III) + 2 H2O
show the reaction diagram
(2)
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
manganese oxidation I
-
-
NIL
-
-
SYSTEMATIC NAME
IUBMB Comments
reactive-black-5:hydrogen-peroxide oxidoreductase
A hemoprotein. This ligninolytic peroxidase combines the substrate-specificity characteristics of the two other ligninolytic peroxidases, EC 1.11.1.13, manganese peroxidase and EC 1.11.1.14, lignin peroxidase. Unlike these two enzymes, it is also able to oxidize phenols, hydroquinones and both low- and high-redox-potential dyes, due to a hybrid molecular architecture that involves multiple binding sites for substrates [2,4].
CAS REGISTRY NUMBER
COMMENTARY hide
114995-15-2
-
42613-30-9
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
strain BOS55, growth without Mn2+ added, stimulation of enzyme synthesis by addition of glycolate, glyoxylate, or oxalate to medium
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
UniProt
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
despite the presence of Mn2+ in the medium, a transformant overexpressing the enzyme produces mnp4 transcripts as well as versatile peroxidase activity as early as 4 days after inoculation. The level of expression is constant throughout 10 days of incubation and the activity is comparable to the typical activity in Mn2+-deficient media
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
1,4-benzohydroquinone + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
1,4-dimethoxybenzene + H2O2
1,4-benzoquinone + H2O
show the reaction diagram
-
Mn2+-independent activity
-
-
?
1-methylanthracene + H2O2
1-methylanthraquinone + H2O
show the reaction diagram
-
at 43% of the rate with 9-methylanthracene
-
-
?
1-naphthol + H2O2
? + H2O
show the reaction diagram
-
-
-
?
2 2,6-dimethoxyphenol + 2 H2O2
coerulignone + 2 H2O
show the reaction diagram
2 2,6-dimethoxyphenol + H2O2
coerulignone + 2 H2O
show the reaction diagram
-
-
-
-
?
2 Mn2+ + 2 H+
2 Mn3+ + H2
show the reaction diagram
-
-
-
-
?
2 Mn2+ + 2 H+ + H2O2
2 Mn3+ + 2 H2O
show the reaction diagram
2 Mn2+ + H2O2 + 2 H+
2 Mn3+ + 2 H2O
show the reaction diagram
-
-
-
-
?
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) + H2O2
? + H2O
show the reaction diagram
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) + H2O2 + H+
? + H2O
show the reaction diagram
-
-
-
?
2,6-dimethoxybenzohydroquinone + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
2,6-dimethoxyphenol + H2O2
?
show the reaction diagram
-
-
-
-
?
2,6-dimethoxyphenol + H2O2
coerulignone + ?
show the reaction diagram
2,7-diaminofluorene + H2O2
? + H2O
show the reaction diagram
-
during oxidation of 2,7-diaminofluorene, both with and without Mn2+, biphasic kinetics with apparent saturation in both micromolar and millimolar ranges are obtained
-
-
?
2-chloro-1,4-dimethoxybenzene + H2O2
2-chloro-1,4-benzoquinone + H2O
show the reaction diagram
-
Mn2+-independent activity
-
-
?
2-methoxy-1,4-benzohydroquinone + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
2-methylanthracene + H2O2
2-methylanthraquinone + H2O
show the reaction diagram
-
at 24% of the rate with 9-methylanthracene
-
-
?
3-hydroxyanthranilic acid + H2O2
? + H2O
show the reaction diagram
-
Mn2+-independent activity
-
-
?
3-methyl-2-benzothiazolinone hydrazone + H2O2
? + H2O
show the reaction diagram
-
enzyme has several substrate binding sites for 3-methyl-2-benzothiazolinone hydrazone, in addition to low and high affinity binding sites for Mn2+
-
-
?
4-aminobenzoic acid + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
4-hydroquinone + H2O2
4-benzoquinone + H2O
show the reaction diagram
-
-
-
-
?
9-methylanthracene + H2O2
10-methylanthracene-9-one + H2O
show the reaction diagram
-
-
-
-
?
acetosyringone + H2O2 + H+
oxidized acetosyringone + H2O
show the reaction diagram
-
-
-
-
?
Acid Blue 62 + H2O2 + H+
oxidized Acid Blue 62 + H2O
show the reaction diagram
-
-
-
-
?
amaranth + H2O2
? + H2O
show the reaction diagram
-
-
-
?
anthracene + H2O2
9,10-anthraquinone + H2O
show the reaction diagram
-
-
-
-
?
anthracene + H2O2
anthraquinone + H2O
show the reaction diagram
-
at 4.8% of the rate with 9-methylanthracene
-
-
?
bovine pancreatic RNase
oxidized bovine pancreatic RNase
show the reaction diagram
-
no redox mediators involved
-
-
?
carbazole + H2O2
? + H2O
show the reaction diagram
-
at 4.8% of the rate with 9-methylanthracene
-
-
?
catechol + H2O2
2-benzoquinone + H2O
show the reaction diagram
-
-
-
-
?
chrysene + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
fluoranthene + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
fluorene + H2O2
9-fluorenone + H2O
show the reaction diagram
-
-
-
-
?
guaiacol + H2O2
3,3'-dimethoxy-4,4'-biphenylquinone + H2O
show the reaction diagram
-
-
-
-
?
guaiacol + H2O2
? + H2O
show the reaction diagram
guaiacol + H2O2 + H+
oxidized guaiacol + H2O
show the reaction diagram
lignin + H2O2
? + H2O
show the reaction diagram
manganese(II)-substituted polyoxometalate + H2O2
manganese(III)-substituted polyoxometalate + H2O2
show the reaction diagram
-
-
-
-
?
methoxyhydroquinone + H2O2
? + H2O
show the reaction diagram
methylene blue + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
Mn2+ + H2O2
?
show the reaction diagram
-
-
-
-
?
Mn2+ + H2O2
Mn3+ + H2O
show the reaction diagram
Mn2+ + H2O2 + 2,6-dimethoxyphenol
?
show the reaction diagram
-
-
-
-
?
Mn2+ + H2O2 + guaiacol
?
show the reaction diagram
-
-
-
-
?
Mn2+ + H2O2 + phenol red
?
show the reaction diagram
-
-
-
-
?
Mn2+ + H2O2 + remazol black-5
?
show the reaction diagram
-
incubation of enzyme with dyes rose bengal, remazol brilliant violet, remazol black-5, remazol blue-19, and remazol orange-16, results in the decolorization of all the dyes tested within a range of 71-84% after 16 h incubation with the enzyme at 100 U/l
-
-
-
Mn2+ + H2O2 + remazol blue-19
?
show the reaction diagram
-
incubation of enzyme with dyes rose bengal, remazol brilliant violet, remazol black-5, remazol blue-19, and remazol orange-16, results in the decolorization of all the dyes tested within a range of 71-84% after 16 h incubation with the enzyme at 100 U/l
-
-
-
Mn2+ + H2O2 + remazol brilliant violet
?
show the reaction diagram
-
incubation of enzyme with dyes rose bengal, remazol brilliant violet, remazol black-5, remazol blue-19, and remazol orange-16, results in the decolorization of all the dyes tested within a range of 71-84% after 16 h incubation with the enzyme at 100 U/l
-
-
-
Mn2+ + H2O2 + remazol orange-16
?
show the reaction diagram
-
incubation of enzyme with dyes rose bengal, remazol brilliant violet, remazol black-5, remazol blue-19, and remazol orange-16, results in the decolorization of all the dyes tested within a range of 71-84% after 16 h incubation with the enzyme at 100 U/l
-
-
-
Mn2+ + H2O2 + rose bengal
?
show the reaction diagram
-
incubation of enzyme with dyes rose bengal, remazol brilliant violet, remazol black-5, remazol blue-19, and remazol orange-16, results in the decolorization of all the dyes tested within a range of 71-84% after 16 h incubation with the enzyme at 100 U/l
-
-
-
Mn2+ + H2O2 + veratryl alcohol
?
show the reaction diagram
-
-
-
-
?
Mordant Black 9 + H2O2 + H+
oxidized Mordant Black 9 + H2O
show the reaction diagram
-
-
-
-
?
NADH + H2O2
NAD+ + H2O
show the reaction diagram
-
-
-
-
?
o-anisidine + H2O2
? + H2O
show the reaction diagram
-
Mn2+-independent activity
-
-
?
Orange II + H2O2
? + H2O
show the reaction diagram
-
-
-
?
p-anisidine + H2O2
? + H2O
show the reaction diagram
-
Mn2+-independent activity
-
-
?
p-dimethoxybenzene + H2O2
benzoquinone + H2O
show the reaction diagram
-
catalyzed by isoforms PS3, PS1
-
-
?
phenanthrene + H2O2
9,10-phenanthrenequinone + H2O
show the reaction diagram
-
-
-
-
?
phenol red + H2O2
oxidized phenol red + H2O
show the reaction diagram
-
Mn2+-dependent activity
-
-
?
Poly R-478
oxidized Poly R-478
show the reaction diagram
-
no redox mediators involved
-
-
?
Poly R-478 + H2O2
?
show the reaction diagram
-
decolorization of the dye
-
-
?
pyrene + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
Reactive Black 5 + H2O2
?
show the reaction diagram
Reactive Black 5 + H2O2
? + H2O
show the reaction diagram
Reactive Black 5 + H2O2
oxidized Reactive Black 5 + H2O
show the reaction diagram
Reactive Blue 38 + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
Reactive Blue 5 + H2O2 + H+
oxidized Reactive Blue 5 + H2O
show the reaction diagram
Reactive Blue 72 + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
Reactive Violet 5 + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
RNase A + H2O2
?
show the reaction diagram
-
-
-
-
?
syringaldazine + H2O2
?
show the reaction diagram
-
-
-
-
?
syringaldehyde + H2O2 + H+
oxidized syringaldehyde + H2O
show the reaction diagram
syringol + H2O2
? + H2O
show the reaction diagram
-
-
-
-
?
vanillylidenacetone + H2O2
? + H2O
show the reaction diagram
-
Mn2+-dependent activity
-
-
?
veratryl alcohol
veratraldehyde + H2O
show the reaction diagram
-
-
-
?
veratryl alcohol + H2O2
veratraldehyde + H2O
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Iron
-
at pH 4.5, Ca2+-depleted enzyme has a high-spin Fe3+
Manganese
Zn2+
-
5 mM, 108% of initial activity
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)
-
substrate inhibition
bovine pancreatic RNase
-
competitive to oxidation of veratryl alcohol, non-competitive to oxidation of Mn2+
-
Co2+
-
5 mM, 80% of initial activity
Cu2+
-
5 mM, 18% of initial activity
Fe2+
-
5 mM, 11% of initial activity
H2O2
-
6.4 mM, 5% loss of activity
Mg2+
-
5 mM, 85% of initial activity
Mn2+
-
above 0.1 mM, severe inhibition of oxidation of veratryl alcohol
N-bromosuccinimide
-
modification of tryptophan residues by N-bromosuccinimide drastically reduces the Mn(II)-independent activity and dye decoloration, while Mn(II)-dependent activity is maintained. Effect is not reversed by addition of mediators
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
70
1-naphthol
pH 2.5, 30C
0.0007 - 2.86
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)
0.01 - 76
2,6-dimethoxyphenol
0.009 - 0.038
2,7-diaminofluorene
0.0059 - 2.24
4-hydroquinone
0.035
acetosyringone
-
pH 5, 25C
0.03
Acid Blue 62
-
pH 5, 25C
0.034 - 10.5
catechol
0.035 - 39.8
guaiacol
0.002 - 5
H2O2
6.4
manganese(II)-substituted polyoxometalate
-
in 0.1 M sodium tartrate, pH 5.0, at 20C
-
0.017 - 3
methoxyhydroquinone
0.007 - 0.013
methylene blue
0.012 - 76.4
Mn2+
0.32
Mordant black 9
-
pH 5, 25C
2.4
p-dimethoxybenzene
-
pH 3.0, isoenzyme PS1
0.008
Phenol red
-
pH 4.5, 30C
0.0013 - 0.022
Reactive Black 5
0.089
Reactive Blue 38
-
pH 4.0
0.04
Reactive Blue 5
-
pH 5, 25C
0.027
Reactive Blue 72
-
pH 4.0
0.047
Reactive Violet 5
-
pH 4.0
0.035 - 0.066
syringaldazine
0.048
syringaldehyde
-
pH 5, 25C
0.2 - 1
syringol
0.005
vanillylidenacetone
-
presence of Mn2+, isoenzyme MP-1, pH 5.0; presence of Mn2+, isoenzyme MP-2, pH 5.0
0.116 - 54.7
veratryl alcohol
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.7 - 365
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)
2.3 - 293
2,6-dimethoxyphenol
0.19 - 0.83
2,7-diaminofluorene
2.4
2-methylanthracene
Bjerkandera adusta
-
pH 4.0
7.2 - 108
4-hydroquinone
8 - 185.6
catechol
9.3 - 185.6
guaiacol
1.8 - 490
H2O2
47
manganese(II)-substituted polyoxometalate
Pleurotus eryngii
-
in 0.1 M sodium tartrate, pH 5.0, at 20C
-
4 - 19
methoxyhydroquinone
0.18 - 2.92
methylene blue
2 - 467
Mn2+
4
p-dimethoxybenzene
Pleurotus eryngii
-
pH 3.0, isoenzyme PS1
0.4 - 11.8
Reactive Black 5
19.8
Reactive Blue 38
Bjerkandera adusta
-
pH 4.0
10
Reactive Blue 72
Bjerkandera adusta
-
pH 4.0
16.9
Reactive Violet 5
Bjerkandera adusta
-
pH 4.0
0.22 - 6.1
syringaldazine
3 - 6
syringol
1.4 - 27.3
veratryl alcohol
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1 - 6480
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)
857
1.3 - 476.2
2,6-dimethoxyphenol
306
21.1 - 21.8
2,7-diaminofluorene
19784
8.9 - 1600
4-hydroquinone
1432
2.4
acetosyringone
Pseudomonas putida
-
pH 5, 25C
16488
240
Acid Blue 62
Pseudomonas putida
-
pH 5, 25C
42603
1 - 70.7
catechol
156
0.6 - 17.6
guaiacol
359
4 - 2650
H2O2
22
7.36
manganese(II)-substituted polyoxometalate
Pleurotus eryngii
-
in 0.1 M sodium tartrate, pH 5.0, at 20C
41296
5.7 - 2850
Mn2+
11
50
Mordant black 9
Pseudomonas putida
-
pH 5, 25C
23397
1600 - 1900
Reactive Black 5
1338
200
Reactive Blue 5
Pseudomonas putida
-
pH 5, 25C
3424
3.3 - 174.3
syringaldazine
819
1.2
syringaldehyde
Pseudomonas putida
-
pH 5, 25C
4943
1.3 - 50.6
veratryl alcohol
471
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2.2
-
substrate Reactive Black 5, pH 3.5, 25C
7.2
-
substrate veratryl alcohol, pH 3.0, 25C
8.4
-
substrate 1,4-benzohydroquinone, pH 5.0, absence of Mn2+
8.8
-
substrate 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), pH 3.5, 25C
9.6
-
substrate 2-methoxy-1,4-benzohydroquinone, pH 5.0, absence of Mn2+
10.8
-
substrate 2,6-dimthoxy-1,4-benzohydroquinone, pH 5.0, absence of Mn2+
26.6
-
pH 4.5, 30C
80
-
pH 4.5
194
-
substrate Mn2+, pH 5.0, 25C
334
-
growth on rich peptone medium containing 0.5 mM Mn2+
559
-
after growth on rich peptone medium
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3 - 3.5
-
lignin peroxidase activity in absence of Mn2+
3.5
-
mutant E37K/V160A/T184M/Q202L
4.2
-
and 3.0, substrate 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), presence of Mn2+
6.5
-
and 3.0, substrate veratryl alcohol, absence of Mn2+; and 3.0, substrate veratryl alcohol, presence of Mn2+; and 5.0, substrate 2,7-diaminofluorene, presence of Mn2+
7
-
and 4.5, substrate 2,7-diaminofluorene, absence of Mn2+; substrate syringaldazine, absence and presence of Mn2+; substrate syringaldazine, absence of Mn2+
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.5 - 6
-
-
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
40
-
lignin peroxidase activity in absence of Mn2+
60
-
manganese peroxidase activity
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.5
-
isoelectric focusing
3.67
-
isoenzyme PS1
3.75
-
isoelectric focusing, isoenzyme MP-2
3.8
-
isoenzyme PS3
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
32000
-
4 * 32000, SDS-PAGE
35244
-
x * 35258, calculated, x * 35244, MALDI-TOF
35258
-
x * 35258, calculated, x * 35244, MALDI-TOF
36400
x * 36400, calculated
40000
-
x * 40000, SDS-PAGE
41000
x * 43000, SDS-PAGE, x * 41000, SDS-PAGE of deglycosylated enzyme
42000
-
x * 45000, isoenzymes PS1, PS2, x * 42000, isoenzyme PS3
45000
-
x * 45000, isoenzymes PS1, PS2, x * 42000, isoenzyme PS3
47000
-
x * 47000, SDS-PAGE
120000
-
gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
tetramer
additional information
-
W170 exposed on enzyme surface is a substrate-binding site both for veratryl alcohol and for polymeric substrates
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
construction of enzyme model and identification of active sites for oxidation of Mn2+ and of aromatic substrates
hanging drop vapor diffusion method, crystal structures of untreated versatile peroxidase (immediately after expression in Escherichia coli and in vitro reconstitution), native versatile peroxidase (treated with Mn2+), D175A variant, and wild-type verstile peroxidase (from Pleurotus eryngii culture)
mutant enzyme W164Y, sitting-drop vapor diffusion method, resolution 1.94 A
-
mutants E140G, P141G, K176G, and E140G/K176G, to 1.6, 2.0, 1.5, 1.7 and 2.35 A resolution, respectively
-
wild-type and mutant M247F
-
resonance Raman and electrochemical study. In solution, enzyme shows a heterogeneous spin population, with the five-coordinated quantum mechanically mixed-spin state being the most populated in the latter. The spin population is sensitively dependent on the pH, temperature, and physical, i.e., solution versus crystal versus immobilized, state of the enzymes. The redox potential for the Fe2+/Fe3+ couple is -260 mV
-
structural changes in the mutants D153H, D153A, R244L, N246H, N246A, D153A/N246A are confined to the distal heme environment
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
35
-
pH 4.5, stable for 1 h
58
-
wild-type, melting temperature
59.4
-
mutant E37K/V160A/T184M/Q202L, melting temperature
79
melting temperature
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, stable
-
4C, pH 4-7, wild-type is stable for 96 h
-
4C, stable for at least 72h
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant protein
resource-Q chromatography
-
Resource-Q column chromatography
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
a recombinant mnp2 construct under the control of Pleurotus ostreatus sdi1 expression signals is introduced into the wild-type Pleurotus ostreatus strain by cotransformation with a carboxin-resistant marker plasmid. Recombinant Pleurotus ostreatus strains with elevated manganese peroxidase (MnP) productivity are successfully isolated. The productivity of the recombinant MnP2 in the present system is not high enough to meet the requirements for industrial applications
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expression in Aspergillus nidulans and Aspergillus niger
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expression in Escherichia coli
expression in Escherichia coli fused to a thioredoxin-hexahistidine tag. Activity of the enzyme increases after removing the tag
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expression in Escherichia coli W3110
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expression in Saccharomyces cerevisiae
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recombinant MnP2 is exclusively expressed and no endogenous MnP isozymes are secreted by the recombinant Pleurotus oseatus srain TM2-18
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
enzyme is the predominantly expressed manganese peroxidase in Mn2+-deficient media, whereas strongly repressed to approximately 1% in Mn2+-supplemented media
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A173R
kcat/KM for Mn2+ is 1.4fold lower than wild-type value, kcat/Km for veratryl alcohol is 1.4fold higher than wild-type value, kcat/Km for Reactive Black 5 is 1.3fold higher than wild-type value
A260F
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kinetics similar to wild-type
A260F/R257A
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site-directed mutagenesis
D175A
kcat/KM for Mn2+ is 842fold lower than wild-type value, kcat/Km for veratryl alcohol is3.2 fold higher than wild-type value, kcat/Km for Reactive Black 5 is 1.8fold higher than wild-type value
E140G
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substitution of bulky residue at the main heme access channel, kinetic analysis
E140G/K176G
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variant attains catalytic efficiencies for oxidation of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonate) at the heme channel similar to those of the exposed tryptophan site W164
E140G/P141G
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substitution of bulky residue at the main heme access channel, kinetic analysis
E140G/P141G/K176G
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variant attains catalytic efficiencies for oxidation of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonate) at the heme channel similar to those of the exposed tryptophan site W164
E140G/W164S/K176G
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variant attains catalytic efficiencies for oxidation of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonate) at the heme channel similar to those of the exposed tryptophan site W164
E36A
kcat/KM for Mn2+ is 258fold lower than wild-type value, kcat/Km for veratryl alcohol is identical to wild-type value, kcat/Km for Reactive Black 5 is 1.2fold higher than wild-type value
E36A/E40A
kcat/KM for Mn2+ is 16000fold lower than wild-type value, kcat/Km for veratryl alcohol is 1.3fold higher than wild-type value, kcat/Km for Reactive Black 5 is 1.1fold higher than wild-type value
E36A/E40A/D175A
kcat for Mn2+ is 149fold lower than wild-type value, kcat/Km for veratryl alcohol is nearly identical to wild-type value, kcat/Km for Reactive Black 5 is 2fold higher than wild-type value
E36A/E40A/D175A/P327ter
kcat for Mn2+ is 149fold lower than wild-type value, kcat/Km for veratryl alcohol is 1.6fold lower than wild-type value, kcat/Km for Reactive Black 5 is 2.4fold higher than wild-type value
E36D
kcat/KM for Mn2+ is 77fold lower than wild-type value, kcat/Km for veratryl alcohol is 1.3fold higher than wild-type value, kcat/Km for Reactive Black 5 is 3.5fold higher than wild-type value
E37K/V160A/T184M/Q202L
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mutant obtained by directed evolution, increase in activity and temperature stability
E40A
kcat/KM for Mn2+ is 1231fold lower than wild-type value, kcat/Km for veratryl alcohol is 1.2fold lower than wild-type value, kcat/Km for Reactive Black 5 is nearly identical to wild-type value
E40D
kcat/KM for Mn2+ is 54fold lower than wild-type value, kcat/Km for veratryl alcohol is 1.3fold lower than wild-type value, kcat/Km for Reactive Black 5 is 2.4fold higher than wild-type value
F142G
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substitution of bulky residue at the main heme access channel, kinetic analysis
H232F
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not involved in long-range electron transfer
K176D
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substitution of bulky residue at the main heme access channel, kinetic analysis
K176G
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substitution of bulky residue at the main heme access channel, kinetic analysis
K215G
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substitution of bulky residue at the main heme access channel, kinetic analysis
K215Q
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substitution of bulky residue at the main heme access channel, kinetic analysis
K264A
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kinetics similar to wild-type
M247F
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92% decrease in efficiency for oxidizing Reactive Black 5
M247L
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kinetics similar to wild-type
N256D/R257D/A260F
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unstable, complete loss of activity
P141G
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substitution of bulky residue at the main heme access channel, kinetic analysis
P76G
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substitution of bulky residue at the main heme access channel, kinetic analysis
P76H
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not involved in long-range electron transfer
R257A/A260F
R257K
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65% decrease in efficiency for oxidizing veratryl alcohol
R257L
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3-fold increase in Km value for veratryl alcohol
S158D
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kinetics similar to wild-type
S158E
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kinetics similar to wild-type
S158E/R257D
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unstable, complete loss of activity
W164S/P76H
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no enzymic activity with veratryl alcohol or Reactive Black 5
W164Y
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site-directed mutagenesis, substitution of Trp-164 by a histidine, serine, or tyrosine residues causes a complete loss of activity on veratryl alcohol and Reactive Black 5
W164Y/R257A/A260F
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site-directed mutagenesis, substitution of Trp-164 by a histidine, serine, or tyrosine residues causes a complete loss of activity on veratryl alcohol and Reactive Black 5
E249D
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no catalytic activity
E249Q
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no catalytic activity
Q266F
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kinetic properties for H2O2 almost identical to those of wild-type, less than half the RNase A-oxidizing activity of wild-type
R263D
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no catalytic activity
R263N
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kinetic properties for H2O2 almost identical to those of wild-type, additional N-glycosylation
V166/168L
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kinetic properties for H2O2 almost identical to those of wild-type
W170A
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kinetic properties for H2O2 almost identical to those of wild-type, no oxidation of veratryl alcohol, decrease in oxodation of RNase A
D153A
mutation minimally affects the second order rate constant for Compound I formation and the specificity constant for H2O2, but substitution dramatically reduces the stability of Compound I
D153A/N246A
mutation reduces the second order rate constant for Compound I formation and the specificity constant for H2O2 less than 30fold, substitution dramatically reduces the stability of Compound I
D153H
mutant is more than an order of magnitude less reactive with H2O2 than wild-type
N246A
mutation inimally affects the second order rate constant for Compound I formation and the specificity constant for H2O2, but substitution dramatically reduces the stability of Compound I
N246H
no detectable peroxidase activity
R244L
mutation abolishes the peroxidase activity, and heme iron of the mutant shows a pH-dependent transition from high spin pH 5 to low spin pH 8.5
additional information
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an engineered N-terminally truncated variant of mutant E37K/V160A/T184M/Q202L displays similar biochemical properties to those of the non-truncated counterpart in terms of kinetics, stability and spectroscopic features. Additional cycles of evolution raised the melting temperature by 8 degrees and significantly increased the enzyme's stability at alkaline pHs. In addition, the Km for H2O2 is enhanced up to 15fold while the catalytic efficiency is maintained, and there is an improvement in peroxide stability
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
using 0.15 M urea, 5 mM Ca2+, 0.02 mM hemin, a 4:1 oxidized-glutathione/reduced-glutathione ratio and 0.1 mg/ml protein at pH 9.5
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
degradation
paper production
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versatile peroxidase reacts with soluble lignin fragments in the absence of added mediators, most probably causing extensive polymerisation of high and intermediate fractions of lignin, and an increase of the small-molecular-mass lignin fraction
synthesis