FIG. 4.
Influence of p6* mutations on expression and maturation of Gag-Pol proteins. H1299 cells were transfected with the indicated provirus plasmids or pcDNA3.1 (mock). (A) For analysis of Gag expression, cells were harvested at 48 h posttransfection, and equal amounts of cell lysates were subjected to immunoblot analysis using a CA-specific antibody. (B and C) To characterize virion-associated Gag and Pol proteins, particles released into cell supernatants were harvested 72 h posttransfection and sedimented through a 20% sucrose cushion. Viral proteins were separated by SDS-PAGE and analyzed by immunoblotting. For detection of Gag-specific products, CA (B1)-specific or MA (B2)-specific monoclonal antibodies or polyclonal sera directed to NC (B3 and B4) or p6gag (B5) were used. Pol-derived products were analyzed using IN (C1)-, RT (C2)-, and PR (C3)-specific polyclonal antisera. The positions of molecular mass markers (in kilodaltons) are shown to the left of the gels, and the arrows to the right of the gels indicate specific protein bands.