This PDF file includes:

• Fig. S1. Tags were placed in locations designed to facilitate labeling and avoid perturbation of RIG-I function.
• Fig. S2. Controls demonstrate that dual labeling of RIG-I does not perturb its function, and FRET reports the distance between hel2i and 2CARD.
• Fig. S3. pppNS ejects 2CARD at high concentrations.
• Fig. S4. ADP-AlF_x subtly but reproducibly increases 2CARD ejection.
• Table S1. RNA sequences used in this study.
• References (50, 51)

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