Recombinant Enterokinase is a high-purity recombinant bovine enterokinase. The enzyme is purified by HPLC with high purity, high specificity and no other proteases. Enterokinase (EC 18.104.22.168) is a specific protease that can cleave the carboxy-terminal site of lysine containing four aspartic acids in the front: Aspartic acid-Aspartic acid-Aspartic acid-Aspartic acid Aspartic acid-lysine. Enterokinase can remove the fusion protein located at the N-terminus of the protein to remove unwanted fusion tags.
1) A specific protease cleaves the carboxy-terminal site of lysine containing four aspartic acids in front: aspartic acid-aspartic acid-aspartic acid-aspartic acid-lysine (DDDDK)
2) No other proteases, no non-specific cutting
Figure 1: SDS-PAGE detection result, a single main band of purified enterokinase
Figure 2: SDS-PAGE analysis of 1U of EK on 50ug substrate 0min, 10min, 20min, 30min, 40min, 50min, 60min
Figure 3: SDS-PAGE analysis of 1U, 2U, 5U, 10U EK on 50ug substrate for 4h and 1U EK on 50ug substrate for 16h
Enzyme digestion conditions:
According to the definition of enzyme activity, an example of recombinant EK digestion conditions: In 25mM Tris-HCl 8.0 system:
Fusion protein concentration 0.1-1mg/ml (total protein 50-100µg)
Reconstituted EK dosage 1-2U
Overnight digestion, or complete digestion requires 12h-16h.
Common factors affecting enterokinase activity
At >200mM imidazole, or >200mMNaCl, or >5% glycerol, the digestion effect will be affected. You can refer to the following recommended methods for digestion:
1) In order to obtain an ideal digestion result, please dialyze the sample into 25mM Tris-HCl 8.0 buffer, and then perform digestion.
2) If it is inconvenient for dialysis, you can dilute the sample. The imidazole content is below 100mM, the NaCl concentration is below 50mM, and the glycerol concentration is below 5% for digestion, and the ratio of enzyme dosage to protein remains unchanged (ie 1U digestion 50μg protein) .
3) If the sample solution contains one or more of the above components and it is inconvenient to remove, at this time, increase the enzyme amount or extend the digestion time to achieve better digestion results.
Used to remove the fusion protein located at the N-terminus of the protein to remove unwanted fusion tags
Source Recombinant Escherichia coli
Theoretical MW: 25,850DA
Activity 1U is defined as storing 50ug at 25℃, within 12h~16h
The amount of enzyme required to cleave 95% of the fusion protein in 25mM Tris-HCl 8.0 buffer.
Storage Store at -20°C.
Stability It can be transported at room temperature and can be stored stably for one week at 25°C.
The enzyme solution can be repeatedly frozen and thawed several times without affecting the enzyme activity.
Product name: Recombinant enterokinase
Specific activity: ≥5u/μl
Packaging: 10mg, 100mg, 1g tube or bottle or bag etc
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