FIG 5.
Binding of murine Ifit1 to the 5′-UTR of IAV. (A) A filter-binding assay was used to measure the binding affinity of murine Ifit1 for the viral RNA (vRNA) of influenza A virus (IAV-Cal). Oligonucleotides representing the first 41 nucleotides of the PB2, PB1, and NS vRNAs were modified to contain a cap 0 (black) or cap 1 (red) structure or left untreated (5′-ppp RNA) (green). Dissociation constants (KD) were calculated by using the nonlinear Hill equation, using Origin. The data are means ± SD and are representative of two independent experiments performed in triplicate. (B) Ifit1 has a lower affinity for RNA with 5′-OH than for 5′-ppp and cap 0 RNAs. A nonradiolabeled RNA (125 nM) representing the 5′ end of PB2 of IAV-Cal was modified to contain a cap 0 or 5′-OH structure and added to the Ifit1 protein (0.3 μM) in the presence of radiolabeled 5′-ppp RNA from PB2 (5 nM). The data represent the means ± SD for two independent experiments performed in triplicate. **, P < 0.02; ns, no significant difference.