The pH dependence of cleavage and folding for wobble-like 8-3 base-pair variants. (A) G8-U3 (black squares), diAP8-C3 (gray squares), and A8-C3 (green squares) (B) pH-dependent folding as monitored by the fluorescence change of pyC7 of wobble-like 8-3 base-pair analogs. The data were acquired, fitted, and plotted as described for G8-C3 folding in Figure 4. Folding of the analyzed variants yielded a bell-shaped pH profile, and fit resulted in two apparent pKas of folding of 5.5 (±0.3) and 6.9 (±0.2) for G8-U3; 6.1 (±0.6) and 6.9 (±0.3) for diAP8-C3; 6.2 (±0.4) and 6.7 (±0.5) for A8-C3. (C) pH-dependent cleavage assays were performed as described for 8-3 analogs in Figure 4. pH dependence of cleavage for each of the analyzed variants yielded a bell-shaped profile, and curve fitting revealed two apparent pKas of 7.9 (±0.4) and >8.5 for G8-U3; 6.7 (±0.4) and 8.1 (±0.3) for diAP8-C3; 6.9 (±0.2) and 8.1 (±0.3) for A8-C3.
