FIG. 4.

GRSF-1 binds the conserved A-box of the influenza virus NP 5′ UTR. Shown here is a representative autoradiogram of RNA gel shift analysis of radiolabeled influenza virus NP (lanes 1 to 7) or NP-A (lanes 8 to 14) 5′ UTR RNA incubated with increasing concentrations (50, 100, and 200 ng) of thrombin-cleaved (rGRSF-1) or uncleaved GST-GRSF-1. Proteins were incubated with radiolabeled RNA (100,000 dpm) in buffer that contained 5 mM HEPES (pH 7.6), 25 mM KCl, 2 mM MgCl₂, 3.8% glycerol, 100 mM NaCl, 0.02 mM dithiothreitol, 2 mM GTP, and 1.5 mM ATP at 30°C for 20 min. Samples were then electrophoresed on a 5% polyacrylamide gel at 4°C and visualized by autoradiography. These data are representative of two independent experiments.