Application | Comment | Organism |
---|---|---|
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 (Comment) | Organism |
---|---|
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 | Comment | Organism |
---|---|---|
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 GaOCBM29. 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] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.008 | - |
locust bean galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
0.037 | - |
guar galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
0.07 | - |
galactoxyloglucan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
0.074 | - |
galactoglucomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
0.076 | - |
galactoxyloglucan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
0.081 | - |
guar galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
0.084 | - |
locust bean galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
0.13 | - |
galactoglucomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
0.14 | - |
galactoxyloglucan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
0.16 | - |
galactoglucomannan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
0.19 | - |
locust bean galactomannan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
0.22 | - |
guar galactomannan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
42.4 | - |
D-galactose | pH 7.0, 30°C, recombinant wild-type enzyme | Fusarium graminearum | |
56 | - |
D-galactose | pH 7.0, 30°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
60.4 | - |
D-galactose | pH 7.0, 30°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Cu2+ | a copper oxidase | Fusarium graminearum | |
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 | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
D-galactose + O2 | Fusarium graminearum | - |
D-galacto-hexodialdose + H2O2 | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Fusarium graminearum | P0CS93 | i.e. Gibberella zeae | - |
Purification (Comment) | Organism |
---|---|
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 | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
D-galactose + O2 | - |
Fusarium graminearum | D-galacto-hexodialdose + H2O2 | - |
? | |
galactoglucomannan + O2 | - |
Fusarium graminearum | ? | - |
? | |
galactoxyloglucan + O2 | - |
Fusarium graminearum | ? | - |
? | |
guar galactomannan + O2 | - |
Fusarium graminearum | ? | - |
? | |
locust bean galactomannan + O2 | - |
Fusarium graminearum | ? | - |
? | |
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 | Comment | Organism |
---|---|---|
GAO | - |
Fusarium graminearum |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
galactose-containing polysaccharides oxidation assay at | Fusarium graminearum |
30 | - |
galactose oxidation assay at | Fusarium graminearum |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
3.25 | - |
galactoglucomannan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
3.33 | - |
galactoglucomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
3.37 | - |
locust bean galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
3.57 | - |
locust bean galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
3.65 | - |
galactoxyloglucan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
3.68 | - |
galactoxyloglucan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
3.98 | - |
galactoglucomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
4.3 | - |
locust bean galactomannan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
4.57 | - |
galactoxyloglucan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
5.18 | - |
guar galactomannan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
5.22 | - |
guar galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
6.7 | - |
guar galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
400 | - |
D-galactose | pH 7.0, 30°C, recombinant wild-type enzyme | Fusarium graminearum | |
516.7 | - |
D-galactose | pH 7.0, 30°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
600 | - |
D-galactose | pH 7.0, 30°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7 | - |
assay at | Fusarium graminearum |
General Information | Comment | Organism |
---|---|---|
additional information | influence of a family 29 carbohydrate binding module on the activity of galactose oxidase from Fusarium graminearum | Fusarium graminearum |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
9.23 | - |
D-galactose | pH 7.0, 30°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
9.43 | - |
D-galactose | pH 7.0, 30°C, recombinant wild-type enzyme | Fusarium graminearum | |
9.93 | - |
D-galactose | pH 7.0, 30°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
20.31 | - |
galactoglucomannan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
22.63 | - |
locust bean galactomannan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
23.02 | - |
galactoxyloglucan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
23.56 | - |
guar galactomannan | pH 7.0, 25°C, recombinant wild-type enzyme | Fusarium graminearum | |
26.07 | - |
galactoxyloglucan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
30.64 | - |
galactoglucomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
45.05 | - |
galactoglucomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
60.09 | - |
galactoxyloglucan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
64.4 | - |
guar galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
75.79 | - |
locust bean galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant CBM29-GaO | Fusarium graminearum | |
181.08 | - |
guar galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum | |
445.8 | - |
locust bean galactomannan | pH 7.0, 25°C, recombinant chimeric enzyme mutant GaO-CBM29 | Fusarium graminearum |