| Background: |
TEV protease, encoded by the tobacco etch virus, is a catalytic domain of the nuclear inclusion a (NIa) protein, consisting of 241 amino acids with a molecular weight of 27 kDa. TEV protease recognizes the amino acid sequence E-X-X-Y-X-Q(or S)/X’, cleaving between Q(or S) and X’, where X and X’ represent any amino acid except X’ cannot be P. The optimal cleavage site is ENLYFQ/G. A major drawback of TEV protease is its tendency to self-cleave at a specific site, resulting in a truncated enzyme with significantly reduced activity. The S219V mutant is approximately 100 times more stable than the wild-type enzyme and is a more efficient catalyst. Due to its absolute specificity and broad operating conditions, such as a wide pH range and ionic strength, TEV protease is more versatile than enzymes like EK and thrombin, particularly in recombinant protein production. The optimal cleavage temperature is 30°C, but the enzyme can function at temperatures as low as 4°C. After digestion, TEV protease can be removed from the reaction using GST chromatography, thanks to its GST tag. |
