FIG. 6.
Specific mutations in eIF4GII have differential effects on 2Apro and Lbpro cleavage. (A) Predicted cleavage activity of 2Apro and Lbpro on eIF4GII mutant proteins. (B) The indicated fusion proteins were incubated for 1 h at 30°C in buffer A alone (lanes 1, 4, and 7), with purified HRV2 2Apro (1.2 μg; lanes 2, 5, and 8), or with purified FMDV Lbpro (1.43 μg; lanes 3, 6, and 9). Protein samples were analyzed by SDS-PAGE and immunoblotting with a monoclonal antibody against the C-terminal FLAG tag (Upstate Biotechnology Inc.). (C) The wt and S701P mutant GST-eIF4GII-FLAG fusion proteins were incubated with the indicated concentrations of purified FMDV Lbpro for 1 h at 30°C. Protein samples were analyzed by SDS-PAGE and immunoblotting as described for panel B.