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TEV Protease

No Product Name Cat.# Conc Size Price(₩) Add
1 TEV Protease PR002S 10 unit/μl 1,000 units ₩ 300,000 추가
2 TEV Protease PR002M 10 unit/μl 2,000 units ₩ 540,000 추가
3 TEV Protease PR002L 10 unit/μl 5,000 units ₩ 1,200,000 추가
Product Name Cat.# Size Price(₩) EA Total(₩) 삭제
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Product description
This product is a recombinant sequence-specific protease purified to near homogeneity from E. coli that expresses a gene encoding TEV Protease of Tobacco Etch Virus. TEV Protease recognizes a 7 amino acid sequence (EXXYXQ ↓ G/S) and cleaves the peptide bond between glutamine and glycine or serine (indicated by an arrow ↓).

Characteristics
- Molecular weight : 28.4 kDa
- Reaction temperature : 4 ~ 30℃
- Optimal pH range : pH 5.5 ~ 8.5 (optimal pH=7)

Applications
- Removal of fusion tag from recombinant proteins

Quality control
- Purity : >95% on SDS-PAGE
- Non-specific protease-free
- Endonuclease-free
- Exonuclease-free
- RNase-free

Unit definition
One unit of TEV Protease is the amount of enzyme required to cleave >85% of 3 μg of substrate protein in 1 hr at 30℃ in the 1X TEV Protease buffer.

Storage buffer
50 mM Tris-HCl (pH 7.5), 1 mM EDTA, 5 mM DTT, 0.1% TritonX-100, 50% glycerol

10X TEV Protease buffer
500 mM Tris-HCl (pH 8.0), 5 mM EDTA, 1 mM DTT

Standard reaction conditions

10X TEV Protease buffer

10 μl

TEV Protease (10 units/μl)

1 μl

Fusion Protein (20 μg/μl)

1 μl

Distilled water

Up to 100 μl

→ Incubate the reaction mixture at 30.

→ Withdraw 20 μl aliquots at 1, 2, 4, and 5 hr after incubation.

→ Add 20 μl of 2X SDS sample buffer to the ailqouts, boil the mixture, and analyze by SDS-PAGE to

    Check cleavage.

→ Keep the rest of the reaction mixture at -20 until analysis is complated

 

- Cleavage efficiency at different lengths of time of incubation and temperatures

Cleavage %

Time

4

16

23

30

0.5 h

37

64

75

87

1 h

58

87

99

99

2 h

76

99

99

97

3 h

89

99

99

99

16 h

96

98

99

99


Cautions
- It is important to confirm cleavage efficiency of TEV Protease with each substrate, since it varies greatly

  depending on the type of substrates used, incubation temperature, incubation period, and the amount of 

  TEV Protease used. Some substrates, despite the presence of a cleavage site, may not be cleaved if the site

  is not accessible to the enzyme. This can be overcome by inserting polyglycine, polyhistidine, or a FLAG-tag

  epitope next to a TEV Protease cleavage site. This allows the recognition sequence to be exposed to the

  TEV Protease.
- After cleavage reaction, TEV Protease can be efficiently removed with a Ni2+-affinity column due to the

  presence of a polyhistidine tag at its N-terminus. Prior to a Ni2+-affinity column, EDTA and DDT should be

  removed. The addition of 10 mM Mg2+ can effectively neutralize EDTA. Generally, substrate proteins bound

  on the affinity resins in a column requires a larger amounts of TEV Protease than proteins in solution.


  

Figure 1. Specific cleavage of substrates by TEV Protease.
Three different substrate proteins (3 μg each) that include MBP-GST, bovine serum albumin (BSA), and mutated MBP-GST (TEV-cleavage site mutant) were incubated at 30℃ for 1 hr with 10 units (1.8 μg) of TEV protease (Cat.# PR002). Only the MBP-GST substrate that contains a correct TEV cleavage site (ENLYFQ^G) present between GST (glutathione S-transferase) and MBP (maltose binding protein) was cleaved into MBP and GST. Neither BSA nor the mutated MBPGST was not cleaved by TEV protease. The mutated MBP-GST contained an amino acid change in the cleavage site. + and - indicate the additions and omissions, respectively, of TEV protease in reaction mixtures. Migration positions of proteins used or products formed were indicated by arrows. Mw, molecular weight marker.



● Material Safety Data Sheet

- Enzynomics_MSDS_PR002_TEV Protease_E

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