Application | Comment | Organism |
---|---|---|
synthesis | application of the engineered enzyme K312E in bioconversion of L-sorbose to 5-keto-D-fructose | Phlebiopsis gigantea |
Cloned (Comment) | Organism |
---|---|
expression in Escherichia coli BL21(DE3), yielding an intracellular and enzymatically active P2OxB with a volumetric yield of 500 units/ml | Phlebiopsis gigantea |
Protein Variants | Comment | Organism |
---|---|---|
E540K | mutant with increased thermo- and pH-stability compared with wild-type, concomitantly with increased catalytic efficiencies (turnover number/KM-value) for D-xylose and L-sorbose | Phlebiopsis gigantea |
K312E | mutant enzyme with significant improvements with respect to the ratio of turnover number to Km-value for D-glucose (5.3fold), methyl-beta-D-glucoside (2.0fold), D-galactose (4.8fold), D-xylose (59.9fold), and L-sorbose (69.9fold) compared with wild-type P2Ox | Phlebiopsis gigantea |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.4 | - |
D-glucose | pH 7.3, 22°C, mutant enzyme K312E with a C-terminal His6-tag | Phlebiopsis gigantea | |
0.6 | - |
D-glucose | pH 7.3, 22°C, mutant enzyme E540K | Phlebiopsis gigantea | |
0.6 | - |
D-glucose | pH 7.3, 22°C, mutant enzyme E540K with a C-terminal His6-tag | Phlebiopsis gigantea | |
0.8 | - |
D-glucose | pH 7.3, 22°C, recombinant wild-type enzyme | Phlebiopsis gigantea | |
1.1 | - |
D-glucose | pH 7.3, 22°C, wild-type enzyme | Phlebiopsis gigantea | |
2.4 | - |
D-xylose | pH 7.3, 22°C, mutant enzyme K312E with a C-terminal His6-tag | Phlebiopsis gigantea | |
2.9 | - |
L-sorbose | pH 7.3, 22°C, mutant enzyme K312E with a C-terminal His6-tag | Phlebiopsis gigantea | |
5 | - |
L-sorbose | pH 7.3, 22°C, mutant enzyme E540K | Phlebiopsis gigantea | |
5.2 | - |
D-galactose | pH 7.3, 22°C, mutant enzyme E540K | Phlebiopsis gigantea | |
5.4 | - |
L-sorbose | pH 7.3, 22°C, mutant enzyme E540K with a C-terminal His6-tag | Phlebiopsis gigantea | |
6.1 | - |
D-galactose | pH 7.3, 22°C, mutant enzyme K312E with a C-terminal His6-tag | Phlebiopsis gigantea | |
6.3 | - |
D-galactose | pH 7.3, 22°C, mutant enzyme E540K with a C-terminal His6-tag | Phlebiopsis gigantea | |
6.6 | - |
D-xylose | pH 7.3, 22°C, mutant enzyme E540K with a C-terminal His6-tag | Phlebiopsis gigantea | |
7.9 | - |
D-xylose | pH 7.3, 22°C, mutant enzyme E540K | Phlebiopsis gigantea | |
8.2 | - |
D-galactose | pH 7.3, 22°C, wild-type enzyme | Phlebiopsis gigantea | |
11.3 | - |
2-deoxy-D-glucose | pH 7.3, 22°C, wild-type enzyme | Phlebiopsis gigantea | |
22.1 | - |
D-allose | pH 7.3, 22°C, mutant enzyme E540K with a C-terminal His6-tag | Phlebiopsis gigantea | |
22.3 | - |
D-allose | pH 7.3, 22°C, mutant enzyme E540K | Phlebiopsis gigantea | |
25 | - |
D-xylose | pH 7.3, 22°C, recombinant wild-type enzyme | Phlebiopsis gigantea | |
29.4 | - |
D-xylose | pH 7.3, 22°C, wild-type enzyme | Phlebiopsis gigantea | |
47 | - |
L-sorbose | pH 7.3, 22°C, recombinant wild-type enzyme | Phlebiopsis gigantea | |
50 | - |
methyl-beta-D-glucoside | pH 7.3, 22°C, mutant enzyme K312E with a C-terminal His6-tag | Phlebiopsis gigantea | |
50 | - |
L-sorbose | pH 7.3, 22°C, wild-type enzyme | Phlebiopsis gigantea | |
55.7 | - |
D-allose | pH 7.3, 22°C, wild-type enzyme | Phlebiopsis gigantea | |
68 | - |
D-allose | pH 7.3, 22°C, mutant enzyme K312E with a C-terminal His6-tag | Phlebiopsis gigantea | |
79 | - |
2-deoxy-D-glucose | pH 7.3, 22°C, mutant enzyme E540K with a C-terminal His6-tag | Phlebiopsis gigantea | |
99.3 | - |
2-deoxy-D-glucose | pH 7.3, 22°C, mutant enzyme K312E with a C-terminal His6-tag | Phlebiopsis gigantea | |
256 | - |
methyl-beta-D-glucoside | pH 7.3, 22°C, mutant enzyme E540K | Phlebiopsis gigantea | |
261 | - |
methyl-beta-D-glucoside | pH 7.3, 22°C, mutant enzyme E540K with a C-terminal His6-tag | Phlebiopsis gigantea | |
289 | - |
methyl-beta-D-glucoside | pH 7.3, 22°C, wild-type enzyme | Phlebiopsis gigantea |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Phlebiopsis gigantea | Q6UG02 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant wild-type and mutant enzymes | Phlebiopsis gigantea |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
2.74 | - |
recombinant wild-type enzyme | Phlebiopsis gigantea |
13.4 | - |
mutant enzyme E540K with a C-terminal His6-tag | Phlebiopsis gigantea |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
2-deoxy-D-glucose + O2 | - |
Phlebiopsis gigantea | 2-deoxy-3-dehydro-D-glucose + H2O2 | - |
r | |
D-allose + O2 | - |
Phlebiopsis gigantea | ? | - |
? | |
D-galactose + O2 | - |
Phlebiopsis gigantea | ? | - |
? | |
D-glucose + O2 | - |
Phlebiopsis gigantea | 2-dehydro-D-glucose + H2O2 | - |
? | |
D-xylose + O2 | - |
Phlebiopsis gigantea | ? | - |
? | |
L-sorbose + O2 | - |
Phlebiopsis gigantea | ? | - |
? | |
methyl-beta-D-glucoside + O2 | - |
Phlebiopsis gigantea | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
P2Ox | - |
Phlebiopsis gigantea |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
45 | - |
wild-type enzyme | Phlebiopsis gigantea |
50 | - |
mutant enzyme K312E with a C-terminal His6-tag | Phlebiopsis gigantea |
Temperature Minimum [°C] | Temperature Maximum [°C] | Comment | Organism |
---|---|---|---|
30 | 58 | 30°C: about 60% of maximal, 58°C: about 70% of maximal activity, mutant enzyme K312E with a C-terminal His6-tag | Phlebiopsis gigantea |
30 | 53 | 30°C: about 60% of maximal activity, 58°C: about 75% of maximal activity | Phlebiopsis gigantea |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
40 | - |
pH 7.0, 30 min, wild-type enzyme, mutant enzyme K312E with a C-terminal His6-tag and mutant enzyme E540K are stable | Phlebiopsis gigantea |
50 | - |
pH 7.0, 30 min, wild-type enzyme loses about 40% of its activity, mutant enzyme K312E with a C-terminal His6-tag and mutant enzyme E540K are stable | Phlebiopsis gigantea |
55 | - |
pH 7.0, 30 min, wild-type enzyme loses 95% of its activity, mutant enzyme K312E with a C-treminal His6-tag and mutant enzyme E540K lose about 10% of its activity | Phlebiopsis gigantea |
60 | - |
pH 7.0, 30 min, wild-type enzyme completely loses its activity, mutant enzyme K312E with a C-terminal His6-tag loses about 40% of its activity and mutant enzyme E540K loses about 10% of its activity | Phlebiopsis gigantea |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
5 | 6 | mutant enzyme E540K | Phlebiopsis gigantea |
6 | - |
mutant enzyme K312E with a C-terminal His6-tag | Phlebiopsis gigantea |
pH Stability | pH Stability Maximum | Comment | Organism |
---|---|---|---|
1 | 2 | pH 7.0, 30 min, wild-type enzyme loses about 75% of its activity, mutant enzyme K312E with a C-terminal His6-tag loses about 70% of its activity and mutant enzyme E540K loses about 10% of its activity | Phlebiopsis gigantea |
6 | - |
30 min, 30 min, wild-type enzyme and mutant enzyme E540K are stable, mutant enzyme K312E with a C-terminal His6-tag loses about 10% of its activity | Phlebiopsis gigantea |
7 | 9 | 30 min, wild-type enzyme, mutant enzyme K312E with a C-terminal His6-tag and mutant enzyme E540K are stable | Phlebiopsis gigantea |
10 | - |
pH 7.0, 30 min, wild-type enzyme loses about 20% of its activity, mutant enzyme K312E with a C-terminal His6-tag and mutant enzyme E540K are stable | Phlebiopsis gigantea |
11 | - |
pH 7.0, 30 min, wild-type enzyme loses about 40% of its activity, mutant enzyme K312E with a C-terminal His6-tag loses about 10% of its activity, mutant enzyme E540K is stable | Phlebiopsis gigantea |