Figure 4
The P1 and P1′ amino acid sites of NS2A/2B are vital for NS2B3 proteolytic processing. (A) Schematic of plasmid construction whereby P1 and/or P1′ were mutated to Alain NS2A/2B-P. (B, C) DEFs were cotransfected with empty vector or a cleavage construct (NS2A/2B-P1, NS2A/2B-P1′ or NS2A/2B-P1P1′) and different concentrations of NS2B3, and the cells were harvested 24 h post transfection for WB analysis. Mouse anti-Flag monoclonal antibody and mouse anti-Myc monoclonal antibodies were used as the primary antibodies simultaneously. (D) NS2A/2B, NS2A/2B-P1, NS2A/2B-P1′ or NS2A/2B-P1P1′ was cotransfected with 400 ng NS2B3 respectively and the cells were harvested 24 h post transfection for WB, Mouse anti-Flag monoclonal antibody and mouse anti-Myc monoclonal antibodies were used as the primary antibodies simultaneously.
