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Merck

T4455

Sigma-Aldrich

TEV Protease

Synonim(y):

P1 protease, TEVp, Tobacco Etch Virus protease, rTEV

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About This Item

Kod UNSPSC:
12352204
NACRES:
NA.54

pochodzenie biologiczne

microbial (Tobacco Etch virus)

Poziom jakości

rekombinowane

expressed in E. coli

opis

Contains both a histidine tag and a GST tag

Postać

solution

aktywność właściwa

≥3,000 units/mg protein

masa cząsteczkowa

27 kDa

metody

protein purification: suitable

przydatność

suitable for purification of HIS tagged recombinant proteins

Zastosowanie

genomic analysis

Warunki transportu

wet ice

temp. przechowywania

−20°C

Opis ogólny

TEV protease has (NIa) protease catalytic domain which corresponds to a molecular weight of 27 kDa. It is unique with high specificity and is active at low temperature.
The tobacco etch virus (TEV) protease is a useful tool for the removal of fusion tags from recombinant proteins.

Zastosowanie

TEV Protease has been used in the removal of histidine tag from recombinant mitogen activated protein kinase 14 (MAPK14), Connexin43 (Cx43) c-terminal fragment and δ1-pyrroline-5-carboxylate reductase from Oryza sativa. It has also been used in the elution of purified proteasomes from human leukemia cell lines K562.

Działania biochem./fizjol.

Immobilized TEV protease on streptavidin-agarose is an efficient tool for fusion protein cleavage. TEV protease has also been used in a study to investigate proteolytic processing of nlrp1b for inflammasome activity.
The tobacco etch virus (TEV) protease undergoes autolysis, but mutants of TEV protease are resistant to autolysis. Bacterial expression of recombinant TEV protease leads to poor yields and less solubility. Use of green fluorescent protein fusion TEV protease overcomes these setbacks and finds application in structural genomics research.
TEV protease has a strict 7 amino acid cleavage recognition sequence of Glu-Asn-Leu-Tyr-Phe-Gln↓Gly/Ser.

Cechy i korzyści

Optimally engineered and purified for enhanced stability and high specific activity over a broad temperature range.

Właściwości fizyczne

A modified 52 kDa TEV protease construct containing both GST and His tags for easy removal using glutathione or His-Select® affinity media.
TEV protease is an engineered catalytic domain of the Tobacco Etch virus NIa protease. TEV protease is a highly specific cystein protease belonging to the C4 peptidase family.

Definicja jednostki

One unit of TEV protease cleaves >85% of 3 μg of control substrate in one hour at pH 8.0 at 30 °C.

Postać fizyczna

Supplied as a >=2 mg/ml in 25 mM Tris-HCl, pH 8.0, 50 mM NaCl, 1 mM TCEP, and 50% glycerol
aqueous glycerol solution

Uwaga dotycząca przygotowania

Cleavage Protocol
Prepare fresh dialysis buffer. Dialysis buffer should be optimized for target protein solubility and contain no protease inhibitors. The dialysis buffer should also be compatible with downstream purification processes, e.g. minimal amount of EDTA or DTT if a HIS Select® column will be used to remove the cleaved His-tag.
Example of suitable dialysis buffer; 25 mM Tris-HCl, pH 8.0, 150 - 500 mM NaCl, 14 mM β-mercaptoethanol
This TEV protease has the same activity in 150 mM NaCl or 500 mM NaCl and 400 mM imidazole.
Dilute the target protein sample to 1-2 mg/ml with dialysis buffer. This is optional in case the target protein aggregates in dialysis buffer. Save a small aliquot as a control for PAGE analysis. EDTA may be added to 0.5 mM final concentration if the target protein will be eluted from the HIS Select column and EDTA is compatible with the target protein.
Add TEV protease at a protease to target protein ratio of 1:100 (w/w) or 10,000 unit (1 mg) TEV protease to 100 mg of target protein. There is no need to calculate the molar ratio. TEV protease can be added directly to the target protein. There is no need to change buffer or dilute TEV protease. The optimal ratio should be determined empirically. A Protease-to-target protein ratio (w/w) of 1:50 to 1:200 should provide an affective range for most target proteins.
Dialyze against the dialysis buffer at 4 °C ~ 16 hrs. Dialysis is intended to remove imidazole or glutathione if HIS Select® or glutathione affinity columns are used to remove the cleaved tag or TEV protease after cleavage.
Typically, 1 mg of TEV protease will cleave >90% of 100mg of a control protein at 4 °C in 16 hours.

Informacje prawne

HIS-Select is a registered trademark of Merck KGaA, Darmstadt, Germany
HiScreen is a trademark of Cytiva
This page may contain text that has been machine translated.

Kod klasy składowania

10 - Combustible liquids

Klasa zagrożenia wodnego (WGK)

WGK 2

Temperatura zapłonu (°F)

Not applicable

Temperatura zapłonu (°C)

Not applicable


Certyfikaty analizy (CoA)

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Masz już ten produkt?

Dokumenty związane z niedawno zakupionymi produktami zostały zamieszczone w Bibliotece dokumentów.

Odwiedź Bibliotekę dokumentów

Functional properties and structural characterization of rice delta1-pyrroline-5-carboxylate reductase
Forlani G, et al.
Frontiers in Plant Science, 6, 565-565 (2015)
Oriented immobilization of the tobacco etch virus protease for the cleavage of fusion proteins
Miladi, B., et al.
Journal of Biotechnology, 128, 97-103 (2012)
Erika Camacho et al.
Biochemical and biophysical research communications, 512(4), 859-863 (2019-04-02)
Abrogation of the hemorrhagic activity of BaP1, a PI Snake Venom Metalloproteinase (SVMP) from the venom of Bothrops asper, was achieved by the substitution of residues in the first part of the Ω loop surrounding the active site by the
Daniel H Pfaff et al.
The Journal of biological chemistry, 292(2), 685-690 (2016-12-03)
Methylglyoxal (MG) is a reactive metabolite that forms adducts on cysteine, lysine and arginine residues of proteins, thereby affecting their function. Methylglyoxal is detoxified by the Glyoxalase system, consisting of two enzymes, Glo1 and Glo2, that act sequentially to convert
Proteomic analysis of affinity-purified extracellular proteasomes reveals exclusively 20S complexes
Kulichkova VA, et al.
Testing, 8(60), 102134-102134 (2017)

Produkty

Read our article about how the Nanodisc system allows for structural studies of membrane proteins.

This page shows how to perform a purification of His-tagged membrane proteins.

Ta strona pokazuje, jak przeprowadzić oczyszczanie białek błonowych znakowanych His.

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