Cloned (Comment) | Organism |
---|---|
gene BPUM_0532, recombinant expression of His-tagged enzyme in Escherichia coli strain Origami B (DE3), evaluation and optimization of a complementary method to optimize the copper oxidase, bilirubin oxidase, enzyme expression in Escherichia coli. Usage of Escherichia coli strain Origami B (DE3), known to promote the formation of disulfide bridges in the bacterial cytoplasm, because the enzyme has a disulfide bridge. In a second step, the effect of coexpression of chaperone proteins (DnaK, DnaJ and GrpE usingthe plasmid pKJE7) on the protein production and specific activity is investigated resulting in an increase of the final amount of enzyme by 858% and its catalytic rate constant by 83%, overview. Comparison of expression efficiency in Escherichia coli strain BL21 (DE3) and Origami B (DE3). When all protein chaperones are expressed together with the r-BOD, the amount of enzyme is largely promoted without impairing the specific activity pointing out the synergy between the two systems, as GroES/GroEL has no effect on the expression alone. Synergy between DnaK, DnaJ, and GrpE, which prevent aggregation of misfolded enzymes, and GroEL and GroES, which promote their proper folding | Bacillus pumilus |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten kinetics | Bacillus pumilus | |
0.032 | - |
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) | recombinant enzyme expressed in Escherichia coli strain BL21(DE3), pH 3.2, 37°C | Bacillus pumilus | |
0.087 | - |
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) | recombinant enzyme expressed in Escherichia coli strain Origami B(DE3), pH 4.0, 37°C | Bacillus pumilus | |
0.124 | - |
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) | recombinant enzyme expressed in Escherichia coli strain BL21(DE3), pH 4.0, 37°C | Bacillus pumilus | |
0.216 | - |
2,6-dimethoxyphenol | recombinant enzyme expressed in Escherichia coli strain BL21(DE3), pH 7.0, 37°C | Bacillus pumilus | |
0.774 | - |
2,6-dimethoxyphenol | recombinant enzyme expressed in Escherichia coli strain Origami B(DE3), pH 7.0, 37°C | Bacillus pumilus | |
0.822 | - |
2,6-dimethoxyphenol | recombinant enzyme expressed in Escherichia coli strain BL21(DE3), pH 6.8, 37°C | Bacillus pumilus |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Cu2+ | the enzyme is a multicopper oxidase | Bacillus pumilus |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Bacillus pumilus | A8FAG9 | - |
- |
Bacillus pumilus SAFR-032 | A8FAG9 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged enzyme from Escherichia coli strain Origami B (DE3) by nickel affinity chromatography and dialysis | Bacillus pumilus |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) + O2 | - |
Bacillus pumilus | ? + H2O | - |
? | |
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) + O2 | - |
Bacillus pumilus SAFR-032 | ? + H2O | - |
? | |
2,6-dimethoxyphenol + O2 | - |
Bacillus pumilus | ? + H2O | - |
? | |
2,6-dimethoxyphenol + O2 | - |
Bacillus pumilus SAFR-032 | ? + H2O | - |
? | |
additional information | usage of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 2,6-dimethoxyphenol as substrates, optimization of the assay method | Bacillus pumilus | ? | - |
? | |
additional information | usage of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 2,6-dimethoxyphenol as substrates, optimization of the assay method | Bacillus pumilus SAFR-032 | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
? | x * 30000, folded recombinant enzyme, SDS-PAGE, x * 60000, unfolded recombinant enzyme, SDS-PAGE | Bacillus pumilus |
Synonyms | Comment | Organism |
---|---|---|
BOD | - |
Bacillus pumilus |
BPUM_0542 | - |
Bacillus pumilus |
copper oxidase | - |
Bacillus pumilus |
MCO | - |
Bacillus pumilus |
multicopper oxidase | - |
Bacillus pumilus |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
37 | - |
assay at | Bacillus pumilus |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
29 | - |
2,6-dimethoxyphenol | recombinant enzyme expressed in Escherichia coli strain BL21(DE3), pH 7.0, 37°C | Bacillus pumilus | |
57 | - |
2,6-dimethoxyphenol | recombinant enzyme expressed in Escherichia coli strain BL21(DE3), pH 6.8, 37°C | Bacillus pumilus | |
100 | - |
2,6-dimethoxyphenol | recombinant enzyme expressed in Escherichia coli strain Origami B(DE3), pH 7.0, 37°C | Bacillus pumilus | |
322 | - |
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) | recombinant enzyme expressed in Escherichia coli strain BL21(DE3), pH 4.0, 37°C | Bacillus pumilus | |
391 | - |
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) | recombinant enzyme expressed in Escherichia coli strain BL21(DE3), pH 3.2, 37°C | Bacillus pumilus | |
716 | - |
2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) | recombinant enzyme expressed in Escherichia coli strain Origami B(DE3), pH 4.0, 37°C | Bacillus pumilus |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
4.2 | 7.2 | assay at | Bacillus pumilus |
pH Minimum | pH Maximum | Comment | Organism |
---|---|---|---|
additional information | - |
inactive at pH 9.2 | Bacillus pumilus |
General Information | Comment | Organism |
---|---|---|
evolution | the enzyme belongs to the family of multicopper oxidases (MCOs) | Bacillus pumilus |