FIG 7.
Effect of BMS-806 and the AEG compounds on the gp120 association with solubilized Env complexes and with Env on VLPs. (A) Triton X-100 lysates of A549-Env cells expressing HIV-1AD8 Env were incubated with Ni-NTA beads in the presence of the indicated compounds at either a 1 or a 10 μM concentration. The precipitated Envs were analyzed by Western blotting with a rabbit anti-gp120 antibody and the 4E10 anti-gp41 antibody. The gp120 association with the Env complex was calculated as described in Materials and Methods for each compound and is shown relative to the value observed for the DMSO control. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, not significant. (B, C) VLPs with the HIV-1AD8 Env were incubated with DMSO, BMS-806, or the AEG compounds in physiological buffer for 4 days at 4°C, or 37°C. The VLPs were then pelleted and lysed; the lysed VLPs and supernatants were analyzed by Western blotting with a rabbit anti-gp120 antibody (top) and anti-Gag p55/p24/p17 antibody (bottom). Note that because the amount of gp120 shed was low compared with the amount of the VLP-associated gp120 shed, the level of gp120 in the VLP supernatants is a more accurate indicator of gp120 shedding. The results of typical experiments from two independent experiments are shown. Lanes M, molecular mass markers. The numbers to the left are molecular masses (in kilodaltons).