Comparative gel electrophoresis of the conformation adopted by the three-way junction of the HCV IRES in different ionic conditions. The junction studied had a simplified helix D, with the central sequence shown in Figure 1C. Three forms of the junction, each with a different helical arm shortened, are electrophoresed in adjacent tracks of a 10% polyacrylamide gel in the presence of 90 mM Tris-borate (pH 8.3) with the indicated metal ions (or EDTA to chelate metal ions). The long–short-arm junction species are named according to the two long arms. An interpretation of the structures adopted is shown on the right, together with the anticipated structures and mobilities of the long–short-arm junction species. In the presence of EDTA the slow-intermediate-fast pattern is interpreted in terms of the extended structure, while the slow-fast-intermediate pattern observed in the presence of metal ions is interpreted in terms of a structure in which helices C and E are coaxially stacked, with an acute angle subtended between helices C and D.
