A region near the C-terminus of FMDV VP1 required for processing of the VP3/VP1 junction.

a) The P1-2A precursor (wt or with small deletions as indicated) was expressed alone or with 3Cpro in transient expression assays in BHK cells as for Fig 3. Cell lysates were prepared and analyzed by immunoblotting. The structural proteins corresponding to the different bands are indicated and were detected using guinea pig anti-FMDV A-Iraq antisera. Bound antibodies were visualized using the anti-guinea pig HRP-conjugated secondary antibodies and chemiluminescence detection. The odd numbered lanes show the P1-2A precursors expressed alone and the even numbered lanes show the precursor co-expressed with 3Cpro. Molecular mass markers (kDa) are indicated on the left. A negative control (No DNA) is included in lane 9. A positive control with a deletion known to be tolerated in replicating FMDV [20] was included, P1-2A (VP1 Δ142–154), lanes 7 and 8. b) The 2A L2P substitution was introduced into the plasmids encoding P1-2A (wt), P1-2A (VP1 Δ191–195), P1-2A (VP1 Δ194–199) and the positive control P1-2A (VP1 Δ142–154). This 2A L2P substitution blocks cleavage of the VP1/2A junction and increased the sensitivity of detection of VP1. The negative control (No DNA) is indicated.