Fig. 4.
After tRNALys3 annealing, NCp7 is required for transition to the extended dimer conformation. Population FRET histograms from FH238.WT dimerized with 238.WT and imaged in 10 mM (A, C, E, and G) or 1 mM (B, D, F, and H) MgCl2 using either the Intra-UTR (Left) or Inter-UTR FRET (Right) assay (Fig. 1), as indicated above for each panel set. Black and red dashed lines indicate the upper and lower bound of low- and high-FRET SD, respectively. Before imaging in both MgCl2 concentrations, the FW238.WT complex was annealed to tRNALys3 (C and D), the 18-nt anti-PBS cDNA (E), or the tRNALys3-derived acceptor-TΨC minihelix (G). Alternatively, the FH238.WT complex was incubated with NCp7 (F) or NCp7 and tRNALys3 (H), followed by removal of NCp7 with proteinase K digestion before imaging (Materials and Methods). The number of FRET trajectories compiled into each histogram is indicated (N).