Figure 3.
Equilibrium binding titrations of fluorescein-labeled ferritin IRE (●), EMCV J/K IRES (γ), β-actin UTR (○), and polyUC (∇) binding to eIF4GI557-1599 (A), eIF4GI682-1599 (B), and DAP5 (C). Briefly, 10 nm (A) or 100 nm (B and C) fluorescein-labeled uncapped RNA oligonucleotides were titrated with increasing concentrations of eIF4GI557-1599, eIF4GI682-1599, or DAP5 in the titration buffer at 25 °C, and the anisotropy at each titration point was measured using excitation and emission wavelengths of 495 nm and 520 nm, respectively. Averages from 3 independent experiments were performed, and the curves represent the nonlinear fits that were used to obtain the averages and standard deviations for the corresponding Kd values given in Table S1.