A Tobacco Etch Virus Protease with Increased Substrate Tolerance at the P1' position
Figure 4
Influence of the P2' residue on substrate degradation.
A) Analysis of tester construct proteolysis and depletion as well as TEV protease production by immunoblotting. Tester constructs (plasmid based): CFP-TDegXY-RFP, XY=RH, RL, X corresponds to the P1' position, Y to the P2' position; proteases: pTEV+ (yeast strain YCT1169), pTEV2 (YCR56). Conditions as in Figure 1B; antibodies directed against tRFP, GFP, TEV, and Tub1 (loading control) were used to obtain the immunoblot. B) The RFP fluorescence of the tester constructs CFP-TDegXY-RFP was followed over time after induction of TEV protease synthesis by fluorimeter measurements (three measurements for each construct; error bars indicate the standard error of the mean; same constructs as in A).