FIG. 3.

Subtilisin treatment of Mo-MuLV virions. COS-7 cells were transiently transfected with plasmids expressing Mo-MuLV and HA-Nuc(212) protein, and virions were purified as for Fig. 2A. Twenty percent of the pellet was digested with the indicated amount of subtilisin, after which PMSF and aprotinin were added to terminate the digestion. (A) Subtilisin digestion in the absence of detergent for 17 h at room temperature, followed by particle purification through 25% sucrose cushion. Thirty percent of the pellet was analyzed by Western blotting. (B) Subtilisin digestion in the presence of 0.2% NP-40 for 1 h at room temperature. Twenty percent of the sample was analyzed by Western blotting. The membranes were probed with a monoclonal anti-HA epitope antibody reprobed with a polyclonal anticapsid serum, and probed again with a monoclonal antienvelope antibody, as indicated on the left. The locations of HA-Nuc(212) and capsid are shown on the right.