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The Preferred TEV Protease for Removing Fusion Protein Tags - Look Here

Time:2023-12-05 Hits:172
Description:
Recombinant TEV Protease (rTEV Protease) has undergone genetic engineering modification and is expressed through recombination in Escherichia coli, carrying a 6X His tag. It not only retains the functional activity of the natural TEV enzyme but also exhibits stronger stability and specificity within a broad temperature range. rTEV Protease is a commonly used tool enzyme for removing label sequences on fusion proteins, with stronger site specificity compared to proteinases like EK and SUMO, and features high activity and specificity.
brand
Item number
Packaging
Seebio®
EBY3644A-1KU
1mL/tube
Seebio®
EBY3644A-10KU
10mL/tube
Seebio®
EBY3644A-50KU
50mL/tube


Features:

It strictly recognizes the seven amino acid sequence EXXYXQ↓(G/S), with the cleavage site between glutamine and glycine/serine. Its sequence specificity is much higher than that of proteinases such as thrombin, factor Xa, and chymotrypsin. TEV Protease can perform enzymatic cleavage within a wide pH range (6.0-8.5) and temperature range (4-30°C), allowing flexibility in reaction conditions based on the target protein. rTEV Protease reaches optimal activity at pH 7.0 and 30°C.


Applications:
Used for removing fusion protein tags from recombinant proteins. The N-terminus of rTEV Protease contains 6×His, and after enzymatic cleavage, rTEV can be conveniently removed through Ni2+ affinity chromatography to achieve the purification of the target protein.


Attributes:

Source: Recombinant expression in Escherichia coli
Appearance: Colorless transparent liquid
Molecular Weight: 27.8 kDa
Specific Activity: 1 U/μL
Unit Definition: The amount of enzyme required to cleave >85% of 3 μg substrate in a 1-hour reaction at 30°C in 1×rTEV Buffer (50 mM Tris, pH 8.0, 0.1 mM EDTA, 1 mM DTT) is defined as one unit of activity.
Purification: Chromatographic purification
Purity: ≥95% (by SDS-PAGE)
Product Composition: 25 mM Tris-HCl, 150 mM NaCl, 1 mM EDTA, 5 mM DTT, 50% (v/v) Glycerol, pH 8.0.


Table 1. rTEV Protease Cleavage Activity at Different Temperatures

Reaction time (h)
Shear activity at different temperatures(%)
4℃
16℃
21℃
30℃
1
34
58
56
70
2
58
80
78
90
3
71
99
99
99
3.5
84
99
99
99


Application examples

Components
Dosage
fusion protein
30μg
rTEV Protease (1 U/μL)
10μL
rTEV buffer(10×)
10μL
ddH2O
Changes with final volume


Prepare the reaction system according to the table above.

Incubate the mixture at 30°C, take samples at 1h, 2h, 4h, and 6h, and place them in separate EP tubes.
Add an appropriate amount of 5×SDS Loading Buffer to the above EP tubes and store at -20°C until cleavage is complete.
Analyze cleaved samples by SDS-PAGE electrophoresis to determine cleavage efficiency and adjust the required enzyme amount and reaction time.
If the target protein is unstable at 30°C, perform the reaction overnight (approximately 16 hours) at 4°C.


Transportation and Storage

Transport at low temperature, store at -20°C.
Shelf life: ≥2 years.


Common Factors Affecting the Activity of Natural TEV Protease

PMSF and AEBSF (1 mM), TLCK (1 mM), Bestatin (1 mg/mL), EDTA (1 mM), PestatinA (1 mM), and E-64 (3 mg/mL). These protease inhibitors do not affect the activity of rTEV.
Zn2+ concentration (>5 mM) has an inhibitory effect on rTEV activity.
Reagents reacting with cysteine are potential inhibitors of rTEV.


Frequently Asked Questions:

Q: What are the advantages of this product?
A: Recombinant TEV Protease (rTEV) is a genetically engineered and purified recombinant protease. It not only exhibits the natural TEV enzyme activity but also avoids the issue of self-cleavage found in natural TEV protease. It demonstrates stronger stability and specificity within a broad temperature range.
Q: What experiments can this product be used for?
A: rTEV is a commonly used tool enzyme for removing affinity tags on fusion proteins and possesses strong site specificity.
Q: What is the optimal cleavage time and temperature for this product?
A: The best reaction time is 3-3.5 hours at a temperature range of 16-30°C.


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For more product details, please contact us: service@seebio.cn or Phone: +86 21 58183719 or Wechat: +86 158 0195 7578