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Literature summary for 1.11.1.13 extracted from

  • Bronikowski, A.; Koschorreck, K.; Urlacher, V.B.
    Redesign of a new manganese peroxidase highly expressed in Pichia pastoris towards a lignin-degrading versatile peroxidase (2018), ChemBioChem, 19, 2481-2489 .
    View publication on PubMed
Search for more literature for 1.11.1.13

Cloned(Commentary)

Cloned (Comment) Organism
codon-optimized gene mrmnp1, high level recombinant expression of wild-type and mutant enzymes in Pichia pastoris strain X-33 with proper heme incorporation in to the resulting enzyme proteins Moniliophthora roreri

Protein Variants

Protein Variants Comment Organism
A172W site-directed mutagenesis, the mutant shows decreased stability compared to the wild-type enzyme in Britton Robinson buffer at pH 3-7 for 24 h measured with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) as a substrate. The mutant shows increased kcat values for all substrates compared to wild-type. The mutant is active with lignin dimers, veratryl alcohol, and the azo dye Reactive Black 5 in contrast to the wild-type enzyme Moniliophthora roreri
A172W/A269R site-directed mutagenesis, the mutant shows decreased stability compared to the wild-type enzyme in Britton Robinson buffer at pH 3-7 for 24 h measured with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) as a substrate. The mutant does not show altered kcat values for all substrates compared to wild-type. The mutant is active with lignin dimers, veratryl alcohol, and the azo dye Reactive Black 5 in contrast to the wild-type enzyme Moniliophthora roreri
A172W/A273T site-directed mutagenesis, the mutant shows decreased stability compared to the wild-type enzyme in Britton Robinson buffer at pH 3-7 for 24 h measured with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) as a substrate. The mutant shows increased kcat values for all substrates compared to wild-type. The mutant is active with lignin dimers, veratryl alcohol, and the azo dye Reactive Black 5 in contrast to the wild-type enzyme Moniliophthora roreri
A172W/F259M site-directed mutagenesis, the mutant shows decreased stability compared to the wild-type enzyme in Britton Robinson buffer at pH 3-7 for 24 h measured with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) as a substrate. The mutant shows increased kcat values for all substrates compared to wild-type. The mutant is active with lignin dimers, veratryl alcohol, and the azo dye Reactive Black 5 in contrast to the wild-type enzyme Moniliophthora roreri
A172W/I171V site-directed mutagenesis, the mutant shows decreased stability compared to the wild-type enzyme in Britton Robinson buffer at pH 3-7 for 24 h measured with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) as a substrate. The mutant shows increased kcat values for all substrates compared to wild-type. The mutant is active with lignin dimers, veratryl alcohol, and the azo dye Reactive Black 5 in contrast to the wild-type enzyme Moniliophthora roreri
A172W/K168V site-directed mutagenesis, the mutant shows increased stability compared to the wild-type enzyme in Britton Robinson buffer at pH 3-7 for 24 h measured with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) as a substrate. The mutant shows increased kcat values for all substrates compared to wild-type. The mutant is active with lignin dimers, veratryl alcohol, and the azo dye Reactive Black 5 in contrast to the wild-type enzyme Moniliophthora roreri
additional information to extend the substrate spectrum of MrMnP1, catalytic tryptophan 172 is introduced at the enzyme surface. Properties of Moniliophthora roreri MrMnP1 manganese peroxidase enzyme are shifted towards those of a versatile peroxidase, comparison with the versatile peroxidase from Pleurotus eryngii (UniProt ID Q9UR19). The resulting mutants demonstrate additional activities towards high-redox-potential substrates, such as lignin dimers, veratryl alcohol, and the azo dye Reactive Black 5. The phenolic and non-phenolic lignin dimers guaiacylglycerol-beta-guaiacyl ether (Ge) and veratrylglycerol-beta-guaiacyl ether (Ve) are tested as substrates at pH 3.0-5.0. The phenolic lignin dimer Ge is barely oxidized by the wild-type enzyme (2% conversion), but after introduction of the A172W mutation around 33% can be degraded at pH 3.0 and pH 4.0, and around 15% at pH 5.0. All additional mutations (except for A269R) further increased the activity towards guaiacylglycerol-beta-guaiacyl ether at pH 3.0 and pH 4.0, with the A172W/K168V mutant showing the highest conversion of up to 56% at pH 3.0. The more recalcitrant non-phenolic lignin dimer veratrylglycerol-beta-guaiacyl ether is oxidized only by the mutants, but not by the wild-type enzyme. The activity of the mutants is more similar to the substrate specificity of EC 1.11.1.14. The wild-type enzyme and the mutants are active with dyes: crystal violet, methyl orange, alizarin red S, Indigo carmine, and remazol brilliant blue R, except for the poor activity of the wild-type enzyme with alizarin red S, overview. The mutants showed similar tendencies, decolorization at pH 3.0 is stronger than that with wild-type enzyme Moniliophthora roreri

Inhibitors

Inhibitors Comment Organism Structure
Mn2+ competitive inhibition Moniliophthora roreri

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information Michaelis-Menten kinetics Moniliophthora roreri
0.0003
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W Moniliophthora roreri
0.0003
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/A269R Moniliophthora roreri
0.0004
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W Moniliophthora roreri
0.0004
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/F259M Moniliophthora roreri
0.0005
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/A269R Moniliophthora roreri
0.0005
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/A273T Moniliophthora roreri
0.0005
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/K168V Moniliophthora roreri
0.0006
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/I171V Moniliophthora roreri
0.0007
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/I171V Moniliophthora roreri
0.001
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/F259M Moniliophthora roreri
0.001
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/K168V Moniliophthora roreri
0.0011
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/A273T Moniliophthora roreri
0.0104
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/A273T Moniliophthora roreri
0.0116
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/A269R Moniliophthora roreri
0.0144
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/F259M Moniliophthora roreri
0.0163
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/K168V Moniliophthora roreri
0.0373
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/I171V Moniliophthora roreri
0.0528
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W Moniliophthora roreri

Localization

Localization Comment Organism GeneOntology No. Textmining
extracellular
-
 Moniliophthora roreri
-
-

Metals/Ions

Metals/Ions Comment Organism Structure
Ca2+ the enzyme binds 2 calcium ions per subunit, required, activates Moniliophthora roreri

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2 Mn(II) + 2 H+ + H2O2 Moniliophthora roreri
-
 2 Mn(III) + 2 H2O
-
 ?
2 Mn(II) + 2 H+ + H2O2 Moniliophthora roreri MCA 2997
-
 2 Mn(III) + 2 H2O
-
 ?

Organism

Organism UniProt Comment Textmining
Moniliophthora roreri V2XS39
-
-
Moniliophthora roreri MCA 2997 V2XS39
-
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2 Mn(II) + 2 H+ + H2O2
-
 Moniliophthora roreri 2 Mn(III) + 2 H2O
-
 ?
2 Mn(II) + 2 H+ + H2O2
-
 Moniliophthora roreri MCA 2997 2 Mn(III) + 2 H2O
-
 ?
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) + 2 H+ + H2O2 substrate of A172W variants mutant enzymes Moniliophthora roreri oxidized 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) + 2 H2O
-
 ?
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) + 2 H+ + H2O2 substrate of A172W variants mutant enzymes Moniliophthora roreri MCA 2997 oxidized 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) + 2 H2O
-
 ?
2,6-dimethoxyphenol + 2 H+ + H2O2 substrate of A172W variants mutant enzymes, very low activity with the wild-type enzyme Moniliophthora roreri oxidized 2,6-dimethoxyphenol + 2 H2O
-
 ?
2,6-dimethoxyphenol + 2 H+ + H2O2 substrate of A172W variants mutant enzymes, very low activity with the wild-type enzyme Moniliophthora roreri MCA 2997 oxidized 2,6-dimethoxyphenol + 2 H2O
-
 ?
alizarin red S + H2O2 substrate of A172W variants mutant enzymes, poor activity with the wild-type enzyme Moniliophthora roreri ? + 2 H2O
-
 ?
crystal violet + H2O2 substrate of wild-type and A172W variants mutant enzymes Moniliophthora roreri ? + 2 H2O
-
 ?
guaiacylglycerol-beta-guaiacyl ether + H2O2 substrate of A172W variants mutant enzymes Moniliophthora roreri ? + 2 H2O
-
 ?
indigo carmine + H2O2 substrate of wild-type and A172W variants mutant enzymes Moniliophthora roreri ? + 2 H2O
-
 ?
methyl orange + H2O2 substrate of wild-type and A172W variants mutant enzymes Moniliophthora roreri ? + 2 H2O
-
 ?
additional information the phenolic and non-phenolic lignin dimers guaiacylglycerol-beta-guaiacyl ether (Ge) and veratrylglycerol-beta-guaiacyl ether (Ve) are tested as substrates at pH 3.0-5.0. The phenolic lignin dimer Ge is barely oxidized by the wild-type enzyme (2% conversion), but after introduction of the A172W mutation around 33% can be degraded at pH 3.0 and pH 4.0, and around 15% at pH 5.0. All additional mutations (except for A269R) further increased the activity towards guaiacylglycerol-beta-guaiacyl ether at pH 3.0 and pH 4.0, with the A172W K168V mutant showing the highest conversion of up to 56% at pH 3.0. The more recalcitrant non-phenolic lignin dimer veratrylglycerol-beta-guaiacyl ether is oxidized only by the mutants, but not by the wild-type enzyme. The activity of the mutants is more similar to the substrate specificity of EC 1.11.1.14. The wild-type enzyme and the mutants are active with dyes: crystal violet, methyl orange, alizarin red S, indigo carmine, and remazol brilliant blue R, except for the poor activity of the wild-type enzyme with alizarin red S, overview. The mutants show similar tendencies, decolorization at pH 3.0 is stronger than that with wild-type enzyme. The wild-type MrMnP1 is able to convert ABTS, 2,6-DMP, and Mn2+, but not high-redox-potential substrates, such as Reactive Black 5 or veratryl alcohol Moniliophthora roreri ?
-
-
additional information the phenolic and non-phenolic lignin dimers guaiacylglycerol-beta-guaiacyl ether (Ge) and veratrylglycerol-beta-guaiacyl ether (Ve) are tested as substrates at pH 3.0-5.0. The phenolic lignin dimer Ge is barely oxidized by the wild-type enzyme (2% conversion), but after introduction of the A172W mutation around 33% can be degraded at pH 3.0 and pH 4.0, and around 15% at pH 5.0. All additional mutations (except for A269R) further increased the activity towards guaiacylglycerol-beta-guaiacyl ether at pH 3.0 and pH 4.0, with the A172W K168V mutant showing the highest conversion of up to 56% at pH 3.0. The more recalcitrant non-phenolic lignin dimer veratrylglycerol-beta-guaiacyl ether is oxidized only by the mutants, but not by the wild-type enzyme. The activity of the mutants is more similar to the substrate specificity of EC 1.11.1.14. The wild-type enzyme and the mutants are active with dyes: crystal violet, methyl orange, alizarin red S, indigo carmine, and remazol brilliant blue R, except for the poor activity of the wild-type enzyme with alizarin red S, overview. The mutants show similar tendencies, decolorization at pH 3.0 is stronger than that with wild-type enzyme. The wild-type MrMnP1 is able to convert ABTS, 2,6-DMP, and Mn2+, but not high-redox-potential substrates, such as Reactive Black 5 or veratryl alcohol Moniliophthora roreri MCA 2997 ?
-
-
Reactive Black 5 + 2 H+ + H2O2 dye decolorization, substrate of A172W variants mutant enzymes Moniliophthora roreri oxidized Reactive Black 5 + 2 H2O
-
 ?
Reactive Black 5 + 2 H+ + H2O2 dye decolorization, substrate of A172W variants mutant enzymes Moniliophthora roreri MCA 2997 oxidized Reactive Black 5 + 2 H2O
-
 ?
remazol brilliant blue R + H2O2 substrate of wild-type and A172W variants mutant enzymes Moniliophthora roreri ? + 2 H2O
-
 ?
veratryl alcohol + H2O2 substrate of A172W variants mutant enzymes Moniliophthora roreri 3,4-dimethoxybenzoic acid + 2 H2O
-
 ?
veratrylglycerol-beta-guaiacyl ether + H2O2 substrate of A172W variants mutant enzymes Moniliophthora roreri ? + 2 H2O
-
 ?

Synonyms

Synonyms Comment Organism
MnP
-
 Moniliophthora roreri
Moror_3885
-
 Moniliophthora roreri
MrMnP1
-
 Moniliophthora roreri
short MnP
-
 Moniliophthora roreri

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
22
-
 assay at room temperature Moniliophthora roreri

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.3
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W Moniliophthora roreri
0.3
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/A269R Moniliophthora roreri
0.4
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/A269R Moniliophthora roreri
0.5
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W Moniliophthora roreri
0.7
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/A273T Moniliophthora roreri
0.7
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/I171V Moniliophthora roreri
0.8
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/F259M Moniliophthora roreri
1
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/K168V Moniliophthora roreri
1.1
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/A273T Moniliophthora roreri
1.1
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/F259M Moniliophthora roreri
1.1
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/I171V Moniliophthora roreri
1.3
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/K168V Moniliophthora roreri
7.9
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/A269R Moniliophthora roreri
8.6
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W Moniliophthora roreri
12.1
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/F259M Moniliophthora roreri
13.4
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/I171V Moniliophthora roreri
16
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/A273T Moniliophthora roreri
16.7
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/K168V Moniliophthora roreri

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
3
-
-
 Moniliophthora roreri

pH Range

pH Minimum pH Maximum Comment Organism
3 8 the highest activity of all enzymes with all tested substrates is at pH 3, except for the WT with 2,6-DMP as a substrate, for which the activity is almost twice as high at pH 8 than that at pH 3. Although the activity of the mutants towards ABTS drops to 5-10% at pH 5 and higher, the residual activities of the mutants towards Reactive Black 5 and veratryl alcohol drops less at higher pH values (20-40% residual activity at pH 5). For 2,6-dimethoxyphenol as a substrate, the activities of MrMnP1 and mutants are lowest at pH 6, but increase at pH 7 and pH 8. The activity at high pH values is transient, presumably due to Ca2+ loss and inactivation of the enzyme. Autoxidation of 2,6-DMP in Britton Robinson buffer at pH 8 in the presence of H2O2 is not detected Moniliophthora roreri

pH Stability

pH Stability pH Stability Maximum Comment Organism
3
-
 recombinant wild-type enzyme retains 26% activity after 24 h, while mutant A172W/K168V shows increased stability and retains 54% activity after 24 h, measured with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) as a substrate Moniliophthora roreri
6
-
 recombinant wild-type enzyme retains 38% activity after 24 h, while mutant A172W/K168V shows increased stability and retains 72% activity after 24 h, measured with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) as a substrate Moniliophthora roreri

Cofactor

Cofactor Comment Organism Structure
heme b the enzyme binds 1 heme b (iron(II)-protoporphyrin IX) group per subunit Moniliophthora roreri

General Information

General Information Comment Organism
evolution maganese peroxidases (MnPs) can be divided into groups of short, long, and extra-long MnPs. The length of long MnPs is 348-361 amino acids, extra-long MnPs are longer than 362 amino acids, and short MnPs are no longer than 348 amino acids. The long and extralong MnPs are characterized by a C-terminal tail that surrounds the main heme access channel, preventing oxidation of low-redox-potential substrates (e.g. ABTS, 2,6-DMP). MrMnP1 is 343 amino acids long and can be categorized as a short MnP Moniliophthora roreri
additional information the catalytic tryptophan 172 is considered to be essential for oxidizing substrates with a high redox potential Moniliophthora roreri

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
9.5
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W Moniliophthora roreri
29.5
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/I171V Moniliophthora roreri
34.5
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/A269R Moniliophthora roreri
55.6
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/F259M Moniliophthora roreri
61.4
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/K168V Moniliophthora roreri
67.3
-
 2,6-dimethoxyphenol pH 7.0, 30°C, mutant A172W/A273T Moniliophthora roreri
116.7
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/I171V Moniliophthora roreri
1000
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W Moniliophthora roreri
1000
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/A269R Moniliophthora roreri
1000
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/A273T Moniliophthora roreri
1100
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/F259M Moniliophthora roreri
1300
-
 veratryl alcohol pH 7.0, 30°C, mutant A172W/K168V Moniliophthora roreri
15800
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/A269R Moniliophthora roreri
19143
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/I171V Moniliophthora roreri
21500
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W Moniliophthora roreri
30250
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/F259M Moniliophthora roreri
32000
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/A273T Moniliophthora roreri
33400
-
 Reactive Black 5 pH 7.0, 30°C, mutant A172W/K168V Moniliophthora roreri