Abstract
Tobacco etch virus protease (TEVp) is a widely used tool enzyme in biological studies. To improve the solubility of recombinant TEVp, three variants, including the double mutant (L56V/S135G), the triple mutant (T17S/N68D/I77V), and the quintuple mutant (T17S/L56V/N68D/I77V/S135G), have been developed, however, with little information on functional stability. Here we investigated the solubility and stability of the three TEVp mutants under different temperature and denaturants, and in Escherichiacoli with different cultural conditions. The quintuple mutant showed the highest solubility and thermostablity, and the double mutant was most resistant to the denaturants. The double mutant folded best in E. coli cells at 37°C with or without the co-expressed molecular chaperones GroEL, GroES and GrpE. The least soluble wild type TEVp displayed better tolerance to denaturants than the triple and the quintuple mutants. All results demonstrated that TEVp is not engineered to embody the most desirable solubility and stability by the current mutations.
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