Targeting the OB-Folds of Replication Protein A with Small Molecules
Figure 2
TDRL-505 inhibits RPA DBD-A/B-DNA interactions. (a) SDS-PAGE analysis of purified RPA DBD-A/B. DBD-A/B was purified as described in “Section 2” and analyzed by electrophoresis on a 12% NuPAGE Bis-Tris Gel. The gel was stained with Coomassie blue. (b) EMSA analysis of TDRL-505 inhibition of DBD-A/B DNA binding activity. Assays were performed as described in “Section 2” with increasing concentrations of TDRL-505 from 10–100 μM in reactions with 12.5 nM DNA and 125 nM DBD-A/B. Products were analyzed by nondenaturing gel electrophoresis, dried, and imaged by phosphorimager analysis. The arrow indicates the position of the free DNA and the bracket the position of the bound DBD-A/B-DNA complex. (c) Quantification of EMSA binding data from Panel (b). The signal representing the RPA-bound and free fractions of DNA was quantified using ImageQuant software, and values represent the mean and SD of triplicate determinations.
