Extended Data Fig. 5: Energetics of base-triple disruption in Tat-ARM binding and cellular transactivation.

a, Changes in fluorescence upon peptide binding is greater for base-triple competent variants than for base-triple disrupted variants. Shown are the fitted minimum and maximum fluorescence values (from equation 1, see Methods) from the TAR-Tat-ARM peptide binding assay for 5 independent experiments. Red dotted lines indicate average maximum values for the base-triple competent variants (190), and base-triple disrupted variants (155). U_0-1 are shown in grey as they are unable to form the base-triple. b, Energy diagram of Tat-ARM peptide binding to base triple competent and base-triple disrupted variants. The peptide can bind a bulge-independent kinked TAR conformation lower in energy than the base-triple disrupted stacked conformation. c, Energy diagram of Tat:SEC binding to TAR in the cellular context. The favorable interactions between Cyclin T1 and the TAR apical loop are unable to form in the kinked state of TAR, and so each base-triple disrupted variant is destabilized by the same amount (c_triple) and binds its non-base triple stacked state (demarcated with an asterisk*). d, Proposed model for an alternative sheared base-triple conformation in the A27U-U38A base-triple disrupting mutants with hydrogen bonds shown as black dashed line (left). Two views of the 3D structural model for the alternative sheared base-triple conformation obtained by replacing A27 with U and U38 with A in the PDBID:6MCE²² U₂ TAR structure (right).