Product Class: Other

TEV Protease
NEBU cloned at NEB recombinant 30 65 Heat

P8112-cutsite

Product Introduction

The TEV protease recognition sequence with the highest catalytic efficiency is ENLYFQ ▼S; however, the amino acid in the P1’ position can also be G, A, M, C, or H (1).

  • Removal of affinity purification tags such as maltose-binding protein (MBP) or poly-histidine from fusion proteins
  • Engineered to prevent autolysis and improve stability
  • Optimal activity and stability for up to 24 months
  • Active in a wide range of buffers; optimal activity between pH 6.0 and 9.0.
  • High substrate specificity with no non-specific proteolysis
  • TEV Protease contains a 7x His-Tag for easy removal from a reaction using NEBExpress® Ni Resin (NEB# S1428S), NEBExpress® Ni Spin Columns (NEB# S1427S/L), or NEBExpress® Ni-NTA Magnetic Beads (NEB# S1423S)
Bioz Badge Exists : True
Catalog # Size Concentration
P8112S 1000.0 units 10000 units/ml

Product Information

Description

TEV Protease is a highly specific cysteine protease. The TEV Protease recognition sequence with the highest catalytic efficiency is ENLYFQ ▼S; however, the amino acid in the P1’ position can also be G, A, M, C, or H (1). It is often used for the removal of affinity purification tags such as maltose-binding protein (MBP) or poly-histidine from fusion proteins. TEV Protease has a 7xHis-tag for easy removal from a reaction using nickel affinity resins and has been engineered to improve thermal stability and decrease autolysis. 

Product Source

Cloned from Tobacco Etch Virus and expressed in E. coli.
This product is related to the following categories:
NEBExpress MBP Fusion and Purification System,
Nickel Purification (His-tag),
Bacterial E. coli Protein Expression Products,
Proteases Products,
Protein Purification Products,
Protein Expression Products,
This product can be used in the following applications:
Fusion Protein Cleavage,
Target Protein Insolubility ,
Protein Purification,
Protein Digestion,
Protein Expression

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  • P8112S     -20    
      TEV Protease P8112SVIAL -20 1 x 0.1 ml 10,000 units/ml
      TEV Protease Reaction Buffer B8035SVIAL -20 1 x 1 ml 10 X

Properties & Usage

Unit Definition

1 unit of TEV Protease will cleave 2 µg of MBP-fusion protein, MBP5-TEV-paramyosin ΔSal, to 95% completion in a total reaction volume of 10 µl in 1 hour at 30°C in 50 mM Tris-HCl (pH 7.5 @ 25°C) with 0.5 mM EDTA and 1 mM DTT.

Reaction Conditions

1X TEV Protease Reaction Buffer
Incubate at 30°C

1X TEV Protease Reaction Buffer
50 mM Tris-HCl
0.5 mM EDTA
1 mM DTT
(pH 7.5 @ 25°C)

Storage Buffer

50 mM Tris-HCl
250 mM NaCl
1 mM TCEP
1 mM EDTA
50% Glycerol
pH 7.5 @ 25°C

Heat Inactivation

65°C for 10 minutes

Molecular Weight

Apparent: 28 kDa

Unit Assay Conditions

Two fold dilutions of TEV Protease are incubated with 2 μg MBP5-TEV-paramyosin ΔSal and 1X TEV Protease Reaction Buffer in a 10 µl reaction. The reaction mix is incubated at 30°C for 1 hour. Separation of reaction products are visualized by SDS-PAGE.

Features

  • Removal of affinity purification tags such as maltose-binding protein (MBP) or poly-histidine from fusion proteins
  • Engineered to improve thermal stability and decrease autolysis
  • High substrate specificity with no non-specific proteolysis 
 

Product Notes

  1. The TEV Protease recognition sequence with the highest catalytic efficiency is ENLYFQ ▼S; however, the amino acid in the P1’ position (S in the recognition sequence) can also be G, A, M, C, or H (1).
  2. If the fusion protein sample contains > 2 M urea, > 0.5 M Guanidine hydrochloride, > 50 mM imidazole, pH values below 6 or above 9, or cysteine protease inhibitors then it will be necessary to dialyze the fusion protein before TEV Protease cleavage.
  3. TEV protease can be used at high concentrations without the occurrence of non-specific proteolysis.
  4. Reactions may be scaled-up linearly to accommodate larger reaction volumes.

References

  1. Kapust, R.B. et al. (2002). Biochem. and Biophysical Research Comm.. 294, 949-955.

Protocols, Manuals & Usage

Protocols

  1. Typical Reaction Conditions for TEV Protease (NEB #P8112)
  2. His-tag removal from protein using TEV Protease

Tools & Resources

Selection Charts

FAQs & Troubleshooting

FAQs

  1. Can TEV Protease recognize and cleave a sequence other than Glu-Asn-Leu-Tyr-Phe-Gln-(Gly/Ser)?
  2. Is it necessary to dialyze the sample prior to cleavage with TEV Protease?
  3. Can TEV Protease be used at lower temperatures?
  4. Is TEV Protease compatible with protease inhibitors?
  5. Is TEV Protease compatible with different reaction buffers?
  6. Can TEV Protease be removed from the reaction after cleavage?

Tech Tips

  • The optimal pH range is 6.0 – 8.0
  • Optimal activity achieved in ≤0.2M NaCl; however, the enzyme retains some activity in up to 2M NaCl
Cleavage at lower temperatures, such as 4ºC, requires overnight incubation

Inhibition occurs in the presence of ≥ 5 mM Zn2+, ≥ 1 mM Cu2+ and ≥ 10 mM Co2+

  • Compatible with 10mM MgSO4, MnCl2 and CaCl2 and up to 100mM EDTA
  • Compatible with the following protease inhibitors: aprotinin, benzamidine, leupeptin, pepstatin, PMSF
  • Can be used at high concentrations with no non-specific proteolysis occurring
  • Some substrates may require extended incubation (up to three days at either 4°C or 30°C)
  • If cleavage is not complete, add more TEV protease after 24 hours and continue incubation

Quality, Safety & Legal

Quality Assurance Statement

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

Certificate Of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Safety DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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