Application | Comment | Organism |
---|---|---|
biotechnology | enteropeptidase is a serine protease used in different biotechnological applications. For many applications the smaller light chain can be used to avoid the expression of the rather large holoenzyme | Homo sapiens |
Cloned (Comment) | Organism |
---|---|
recombinant expression of enteropeptidase light chain as thioredoxin-fusion protein in Escherichia coli strain BL21(DE3) in aggregated form, subcloning in Escherichia coli strain DH5alpha | Homo sapiens |
Protein Variants | Comment | Organism |
---|---|---|
C112S | site-directed mutagenesis, replacement of the free cysteine residue with serine improves the refolding yield of the recombinant enzyme by 50%. The heat stability of this C112S variant was also significantly improved by supercharging | Homo sapiens |
additional information | even mild protein surface supercharging by mutagenesis can have pronounced effects on protein solubility and stability, overview | Homo sapiens |
N6D/G21D/G22D/N141D/K209E | site-directed mutagenesis, the mutations lead to supercharging of the protein surface leading to 100fold increased protein solubility | Homo sapiens |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Homo sapiens | P98073 | - |
- |
Purification (Comment) | Organism |
---|---|
refolded, soluble, and detagged recombinant enteropeptidase light chain by affinity chromatography on soybean trypsin inhibitor agarose | Homo sapiens |
Renatured (Comment) | Organism |
---|---|
recombinant expression of thioredoxin-fusion enteropeptidase light chain from aggregates in 3 M guanidine-HCl, pH 8.0, in Escherichia coli strain BL21(DE3) by dilution of the protein in 0.7 M arginine-HCl, pH 8.5, 15% v/v glycerol, 3 mM reduced glutathione, and 1 mM EDTA. After 72 h at 4°C the refolding solution is dialyzed for 8 h against 50 mM Tris-HCl, pH 8.0, to facilitate complete autocatalytic activation by cleaving the fusion tag | Homo sapiens |
Source Tissue | Comment | Organism | Textmining |
---|---|---|---|
commercial preparation | - |
Homo sapiens | - |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
22 | - |
assay at | Homo sapiens |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
60 | - |
purified recombinant enteropeptidase light chain wild-type and N6D/G21D/G22D/N141D/K209E mutant enteropeptidase light chains show 60% remaining activity | Homo sapiens |
65 | - |
purified recombinant enteropeptidase light chain wild-type and mutant C112S show 10% remaining activity, the purified recombinant N6D/G21D/G22D/N141D/K209E mutant enteropeptidase light chain shows 20% activity remaining | Homo sapiens |
70 | - |
purified recombinant enteropeptidase light chain wild-type and mutant C112S are inactivated, purified recombinant N6D/G21D/G22D/N141D/K209E mutant enteropeptidase light chain shows 20% activity remaining | Homo sapiens |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | - |
assay at | Homo sapiens |