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Fig. 3. NoLS possesses a stronger affinity for B23 than does HIV-1 Rev. Pull-down experiments were performed using GST, GST-NoLS and GST-Rev on the same whole HeLa cell extract for increasing incubation times (1.5, 3 and 20 hours) and the binding of B23 was analyzed. (A) A sample (1.5%) of the proteins eluted after each pull-down assay was electrophoresed and analyzed by Coomassie Blue staining. No significant loss of GST, GST-NoLS or GST-Rev occurred during the pull-down assays. (B) A sample (1.5%) of the extract used for each pull-down assay (Input) and 1.5% of the proteins eluted after the indicated pull-down assay were electrophoresed and immunoblotted using anti-B23 antibodies. The signals corresponding to B23 were quantified and are expressed as a percentage of B23 present in the Input. By contrast to GST, both GST-NoLS and GST-Rev bound B23, but B23-NoLS interaction occurred more rapidly and more efficiently than B23-Rev interaction.

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