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Literature summary extracted from

  • Mollerup, F.; Parikka, K.; Vuong, T.V.; Tenkanen, M.; Master, E.
    Influence of a family 29 carbohydrate binding module on the activity of galactose oxidase from Fusarium graminearum (2016), Biochim. Biophys. Acta, 1860, 354-362 .
    View publication on PubMed

Application

EC Number Application Comment Organism
1.1.3.9 synthesis galactose oxidase (GaO) selectively oxidizes the primary hydroxyl of galactose to a carbonyl, facilitating targeted chemical derivatization of galactose-containing polysaccharides, leading to renewable polymers with tailored physical and chemical properties. Increased substrate binding impeded the action of GaO fusions on more concentrated preparations of galactomannan, galactoglucomannan, and galactoxyloglucan Fusarium graminearum

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.1.3.9 gene gao, recombinant expression of wild-type enzyme and chimeric enzyme mutants GaO-CBM29 and CBM29-GaO in Pichia pastoris strain KM71H Fusarium graminearum

Protein Variants

EC Number Protein Variants Comment Organism
1.1.3.9 additional information a family 29 glucomannan binding module, CBM29-1-2, from Piromyces equi is separately linked to the N- and C-termini of GaO, effects on enzyme activity and binding of GaO towards various polysaccharides. The chimeric enzyme mutants demonstrate enhanced binding to galactomannan, galactoglucomannan and galactoxyloglucan compared to the wild-type enzyme. The position of the CBM29 fusion affects the enzyme function. Particularly, C-terminal fusion leads to greatest increases in galactomannan binding and catalytic efficiency, where relative to wild-type GaO, kcat/Km values increases by 7.5 and 19.8times on guar galactomannan and locust bean galactomannan, respectively. The fusion of CBM29 also induces oligomerization of GaO–CBM29. Removing CBM32 from wild-type GaO leads to complete loss in enzyme activity, and substituting the native CBM32 for CBM29-1-2 does not regain GaO function Fusarium graminearum

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.1.3.9 0.008
-
 locust bean galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 0.037
-
 guar galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 0.07
-
 galactoxyloglucan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 0.074
-
 galactoglucomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 0.076
-
 galactoxyloglucan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 0.081
-
 guar galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 0.084
-
 locust bean galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 0.13
-
 galactoglucomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 0.14
-
 galactoxyloglucan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 0.16
-
 galactoglucomannan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 0.19
-
 locust bean galactomannan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 0.22
-
 guar galactomannan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 42.4
-
 D-galactose pH 7.0, 30°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 56
-
 D-galactose pH 7.0, 30°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 60.4
-
 D-galactose pH 7.0, 30°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
1.1.3.9 Cu2+ a copper oxidase Fusarium graminearum
1.1.3.9 additional information addition of copper sulfate to shake-flask cultivations doubles mutant GaO-CBM29 activity, whereas enzyme production in a bioreactor system increases the yield of CBM29-GaO and GaO-CBM29 by more than 12times and 6times, respectively. Corresponding specific activities also increase by more than 20%. Addition of 0.5 mM copper (II) sulfate to reaction mixtures containing purified GaO, CBM29-GaO, and GaO-CBM29, or treatment with potassium ferricyanide, does not increase corresponding specific activities, confirming that the purified proteins are in the fully oxidized and active state Fusarium graminearum

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.1.3.9 D-galactose + O2 Fusarium graminearum
-
 D-galacto-hexodialdose + H2O2
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.1.3.9 Fusarium graminearum P0CS93 i.e. Gibberella zeae
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.1.3.9 recombinant wild-type enzyme and chimeric enzyme mutants GaO-CBM29 and CBM29-GaO from Pichia pastoris strain KM71H by nickel affinity and hydrophobic interaction chromatography or vice-versa, follwed by dialysis and anion exchange chromatography Fusarium graminearum

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.1.3.9 D-galactose + O2
-
 Fusarium graminearum D-galacto-hexodialdose + H2O2
-
 ?
1.1.3.9 galactoglucomannan + O2
-
 Fusarium graminearum ?
-
 ?
1.1.3.9 galactoxyloglucan + O2
-
 Fusarium graminearum ?
-
 ?
1.1.3.9 guar galactomannan + O2
-
 Fusarium graminearum ?
-
 ?
1.1.3.9 locust bean galactomannan + O2
-
 Fusarium graminearum ?
-
 ?
1.1.3.9 additional information galactose oxidase (GaO) selectively oxidizes the primary hydroxyl of galactose to a carbonyl, facilitating targeted chemical derivatization of galactose-containing polysaccharides, leading to renewable polymers with tailored physical and chemical properties. The activity of wild-type GaO and GaO fusions is measured using the chromogenic ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)) assay Fusarium graminearum ?
-
 ?

Synonyms

EC Number Synonyms Comment Organism
1.1.3.9 GAO
-
 Fusarium graminearum

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.1.3.9 25
-
 galactose-containing polysaccharides oxidation assay at Fusarium graminearum
1.1.3.9 30
-
 galactose oxidation assay at Fusarium graminearum

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.1.3.9 3.25
-
 galactoglucomannan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 3.33
-
 galactoglucomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 3.37
-
 locust bean galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 3.57
-
 locust bean galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 3.65
-
 galactoxyloglucan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 3.68
-
 galactoxyloglucan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 3.98
-
 galactoglucomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 4.3
-
 locust bean galactomannan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 4.57
-
 galactoxyloglucan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 5.18
-
 guar galactomannan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 5.22
-
 guar galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 6.7
-
 guar galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 400
-
 D-galactose pH 7.0, 30°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 516.7
-
 D-galactose pH 7.0, 30°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 600
-
 D-galactose pH 7.0, 30°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.1.3.9 7
-
 assay at Fusarium graminearum

General Information

EC Number General Information Comment Organism
1.1.3.9 additional information influence of a family 29 carbohydrate binding module on the activity of galactose oxidase from Fusarium graminearum Fusarium graminearum

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.1.3.9 9.23
-
 D-galactose pH 7.0, 30°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 9.43
-
 D-galactose pH 7.0, 30°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 9.93
-
 D-galactose pH 7.0, 30°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 20.31
-
 galactoglucomannan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 22.63
-
 locust bean galactomannan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 23.02
-
 galactoxyloglucan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 23.56
-
 guar galactomannan pH 7.0, 25°C, recombinant wild-type enzyme Fusarium graminearum
1.1.3.9 26.07
-
 galactoxyloglucan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 30.64
-
 galactoglucomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 45.05
-
 galactoglucomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 60.09
-
 galactoxyloglucan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 64.4
-
 guar galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 75.79
-
 locust bean galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO Fusarium graminearum
1.1.3.9 181.08
-
 guar galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum
1.1.3.9 445.8
-
 locust bean galactomannan pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 Fusarium graminearum