Tobacco Etch Virus Proteinase, mutant (mTEVP, Recombinant) is used as a valuable tool for cleaving affinity tags and fusion proteins in recombinant protein purification protocols. The wild type TEVP contains a self-cleavage site that the protein recognizes and cleaves. TEV Protease, Recombinant (rTEV) is a site-specific protease purified from E. coli by the affinity tag, GST tag. The protease can be used for the removal of affinity tags from fusion proteins. Following digestion, TEV Protease can be removed from the reaction via the GST tag sequence by affinity chromatography.
The seven-amino-acid recognition site for rTEV is Glu-Asn-Leu-Tyr-Phe-Gln-Gly with cleavage occurring between Gln and Gly.
Recommended Conditions for Cleavage of a Fusion Protein:
A number of variables can be changed to optimize the cleavage of any specific protein. The amount of rTEV, the temperature of the incubation, and the time needed for cleavage may be examined. If the protein of interest is heat-labile, then 4 degrees C incubations are recommended. Reactions at 4 degrees C will require longer incubation times and/or more rTEV.
One unit of rTEV cleaves (same/more than)85% of 3ug control substrate in 1hr at 30 degrees C.
Optimal Reaction Temp:
The optimal temperature for cleavage is 30 degrees C; however, the enzyme can be used at temperatures as low as 4 degrees C.
Storage and Stability:
Aliquot to avoid repeated freezing and thawing and freeze at -70 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Aliquots are stable for 6 months.