Fig. 2

Mutational analysis of the MVEV NS1-2A cleavage site. COS-7 cells were transfected with pNS1-2A.HA wt plasmid DNA or mutant constructs with amino acid substitutions in the octapeptide motif or at the P1’ residue. At 2 days post-transfection, the cells were metabolically labelled for 0.5 h and chased for 0.5 h. Cell lysates were subjected to immunoprecipitation with an anti-HA tag mAb and proteins separated by SDS-PAGE (12.5 % acrylamide). Mutations introduced at the NS1-2A cleavage site are indicated at the top of each panel and % cleavage is shown under each lane. Position and size (in kilodaltons) of marker proteins are indicated on left, and bands corresponding to NS1-2A precursors and cleaved NS2A are denoted by a bracket and arrow, respectively. Arrowheads indicate a putative NS2A related product (NS2A*) of larger size than authentic NS2A. Data are representative of at least 3 independent experiments