FIG 4.
Binding of HIV-1 RT-conjugated magnetic beads with the WT or the indicated substitutions within the context of full-length IN or IN220–270. (A) Full-length IN. Bound and unbound fractions were obtained as described in Materials and Methods. The final volume of the unbound fraction was 500 μl, and a 40-μl aliquot was analyzed by SDS-PAGE and immunoblotting. The level of WT IN binding to RT was normalized to 100%, and the values of mutant INs are expressed as percentages of the binding of the WT IN. Data are expressed as the means ± standard deviations, and the numbers in parentheses represent the number of experiments. Asterisks denote significant differences from WT (P ≤ 0.01). (B) IN220–270. The amounts of IN in the input, bound, and unbound fractions are denoted I, B, and U, respectively. The unbound fraction was TCA precipitated, and the pellet was resuspended in 30 μl of loading buffer before analysis by SDS-PAGE and immunoblotting.