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Literature summary extracted from

  • Simeonov, P.; Berger-Hoffmann, R.; Hoffmann, R.; Straeter, N.; Zuchner, T.
    Surface supercharged human enteropeptidase light chain shows improved solubility and refolding yield (2011), Protein Eng. Des. Sel., 24, 261-268.
    View publication on PubMed

Application

EC Number Application Comment Organism
3.4.21.9 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(Commentary)

EC Number Cloned (Comment) Organism
3.4.21.9 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

EC Number Protein Variants Comment Organism
3.4.21.9 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
3.4.21.9 additional information even mild protein surface supercharging by mutagenesis can have pronounced effects on protein solubility and stability, overview Homo sapiens
3.4.21.9 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

EC Number Organism UniProt Comment Textmining
3.4.21.9 Homo sapiens P98073
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.4.21.9 refolded, soluble, and detagged recombinant enteropeptidase light chain by affinity chromatography on soybean trypsin inhibitor agarose Homo sapiens

Renatured (Commentary)

EC Number Renatured (Comment) Organism
3.4.21.9 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

EC Number Source Tissue Comment Organism Textmining
3.4.21.9 commercial preparation
-
 Homo sapiens
-

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.4.21.9 22
-
 assay at Homo sapiens

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
3.4.21.9 60
-
 purified recombinant enteropeptidase light chain wild-type and N6D/G21D/G22D/N141D/K209E mutant enteropeptidase light chains show 60% remaining activity Homo sapiens
3.4.21.9 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
3.4.21.9 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

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.4.21.9 8
-
 assay at Homo sapiens