Fig. 7.

HIV-1 mature capsid disassembly and in vitro assembly assays. (A) In vitro assembly reaction with purified HIV-1 CA. Purified HIV-1 CA protein (100 μM) was incubated with various doses of BMMP at 37°C for 60 min in buffer at high salt concentration. All samples included 1% DMSO. Assembly products were recovered by centrifugation and subjected to SDS-PAGE followed by Coomassie brilliant blue staining. The band intensities were semiquantified by ImageJ software. For quantification, the pelleted products were subjected to p24CA antigen capture ELISA. Data were shown as means with standard deviations from 3 independent experiments. *, P < 0.05. Assembly products were also negatively stained and examined by electron microscopy. Micrographs were shown at the same magnification. Bar = 100 nm. (B) Cell-free assays for HIV-1 uncoating. HIV-1 mature capsids were isolated through a 1% Triton X-100 layer as described previously (3). The HIV-1 cores were incubated with various doses of BMMP at 37°C up to 120 min. For comparison, the cores were similarly incubated with 50 μM AZT. All samples included 1% DMSO. Residual intact cores were recovered by ultracentrifugation and quantified by p24CA antigen capture ELISA. Data were shown as means with standard deviations from 4 independent experiments. *, P < 0.05; **, P < 0.01.