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. Author manuscript; available in PMC: 2019 Mar 26.
Published in final edited form as: Mol Cell. 2018 Sep 27;72(2):355–368.e4. doi: 10.1016/j.molcel.2018.08.021

Figure 4. Directionality of RIG-I translocation and role of Helicase motif IVa.

Figure 4.

(A) Biotin-streptavidin displacement assay schematic. (B) Time-course of biotin-streptavidin displacement from 5’ppp dsRNA (25 nM) by RIG-I (25 nM). (C) 27bp methylated patch 5’ppp dsRNAs to assess directionality of RIG-I translocation (Red bars- 5nt patch of 2’-O-methylated nucleotides placed 15bp downstream of the 5’ppp end). (D) Streptavidin displacement activity on methylated patch 5’ppp dsRNAs. Error bars are SEM from triplicates. (E) Helicase motif IVa of RIG-I (red) and MDA5 (blue). T667E and T671E in RIG-I are shown as yellow circles. (F) Streptavidin displacement by motif IVa mutants on the 5’ppp dsRNA. Error bars are SEM from triplicates. (G) IFN-β reporter activity of RIGI transfected HEK293T cells upon activation by the 2’-O-methylated 5’ppp dsRNAs. Error bars are SEM from quadruplicates. See also Figure S4.