Activating Compound | Comment | Organism | Structure |
---|---|---|---|
Detergents | the mutant enzyme AmyUS100DELTAIG/M197A is found to be very stable in the presence of solid detergents (commercial detergents). The activity is increased in somes cases. Incubation of the enzyme at 40°C with Ariel, Skip, Dixan, Det, NewDet, Nadhif,m and OMO increases the activity between 10 and 50%. The activity of the engineered enzyme persists at 60°C of the majority of the commercial solid detergents. The incubation with the liquid detergents Lav+ and Nadhif increases the activity by 10-20%, respectively. The presence of the Dinol detergent, the activity decreases to 97 and 91% at 40 and 60°C, respectively | Geobacillus stearothermophilus |
Application | Comment | Organism |
---|---|---|
detergent | AmyUS100DELTAIG is designed to improve the thermostability of the thermoactive and thermostable maltohexaose forming alpha-amylase produced in Geobacillus stearothermophilus sp. US100, AmyUS100 | Geobacillus stearothermophilus |
Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli ER2566 | Geobacillus stearothermophilus |
Protein Variants | Comment | Organism |
---|---|---|
M197A | site-directed mutagenesis. Studies of its catalytic properties show no effect on the thermostability, pH activity/stability, calcium demand and chelator resistance. Specific activity is decreased from 1000 to 845 U/mg. The profile of starch hydrolysis is affected. As a result hereof AmyUS100DELTAIG/M197A produces in majority maltose and maltotriose as major products compared to maltohexaose and maltopentaose produced by the wild-type and the AmyUS100DELTAIG variant. The mutant retains 85% of its original activity. 70% of the mutantM197A activity is retained after 60 min of treatment at 60°C in the presence of 1.8 M H2O2, whereas AmyUS100DELTAIG is totally inactivated. These results confirm the importance of Met197 in the oxidative sensibility, situated in the cavity of the active site | Geobacillus stearothermophilus |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
EDTA | AmyUS100DELTAG retains 40 and 25% of its original activity at 40 and 60°C, respectively, when incubated with 200 mM of EDTA, compared to 12 and 0% for AmyUS100 | Geobacillus stearothermophilus | |
linear alkylbenzene sulfonate | mutant M197A, concentration of 10% incubation at 60°C for 1 h, 71% residual relative activity, assay at 80°C and pH 5.6 | Geobacillus stearothermophilus | |
SDS | mutant M197A, concentration of 10% incubation at 60°C for 1 h, 45% residual relative activity, assay at 80°C and pH 5.6 | Geobacillus stearothermophilus | |
Triton X-100 | mutant M197A, concentration of 10% incubation at 60°C for 1 h, 71% residual relative activity, assay at 80°C and pH 5.6 | Geobacillus stearothermophilus | |
Tween-20 | mutant M197A, concentration of 10% incubation at 60°C for 1 h, 94% residual relative activity, assay at 80°C and pH 5.6 | Geobacillus stearothermophilus | |
Tween-80 | mutant M197A, concentration of 10% incubation at 60°C for 1 h, 88% residual relative activity, assay at 80°C and pH 5.6 | Geobacillus stearothermophilus |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Geobacillus stearothermophilus | P06279 | sp. US100 | - |
Oxidation Stability | Organism |
---|---|
activity of AmyUS100DELTAIG and Termamyl300 (commercial amylase used in detergent) are instantaneously decreased in the presence of 1.8 M H2O2, whereas AmyUSDELTAIG/M197A retains 85% of its original activity. 70% of the mutantM197A activity is retained after 60 min of treatment at 60°C. Termamyl300 retains 20% of its original activity and AmyUS100DELTAIG is totally inactivated. These results confirm the importance of Met197 in the oxidative sensibility, situated in the cavity of the active site | Geobacillus stearothermophilus |
Purification (Comment) | Organism |
---|---|
recombinant protein | Geobacillus stearothermophilus |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
additional information | - |
decreased from 1000 to 845 U/mg for AmyUS100DELTAIG to the mutant M197A, assay at 80°C and pH 5.6 | Geobacillus stearothermophilus |
Storage Stability | Organism |
---|---|
spry-dried enzyme, after 6 months in liquid detergent, 90% of its original activity retained. After 1 year the activity decreases to 61%. The activity of the enzyme in solid detergent is almost invariant. In presence of commercial proteases salvinase and alcalase the enzyme activity decreases by 22 and 18%, respectively, after 6 months | Geobacillus stearothermophilus |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
starch + H2O | - |
Geobacillus stearothermophilus | maltohexaose + ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
AmyUS100 | variant of the most thermoactive and thermostable maltohexaose forming alpha-amylase produced in Geobacillus stearothermophilus sp. US100 | Geobacillus stearothermophilus |
AmyUS100DELTAIG | recombinant protein | Geobacillus stearothermophilus |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
80 | - |
assay at | Geobacillus stearothermophilus |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
5.6 | - |
assay at | Geobacillus stearothermophilus |
pH Minimum | pH Maximum | Comment | Organism |
---|---|---|---|
4 | 9 | AmyUS100 and mutant M197A enzyme | Geobacillus stearothermophilus |