RNAs used to assess the effects of RNA secondary structure on PPR10 binding.

(A) Sequences and predicted secondary structures of the RNA ligands. PPR10 is shown aligned to its 23-nt in vivo footprint near atpH (atpH-23mer). PPR10’s minimal binding site is underlined [9]. The atpH-23mer is not predicted to form any structure. Nucleotides that are appended to the PPR10 footprint to introduce RNA structure are colored. (B) Predicted and measured stabilities of each RNA structure at 1 M NaCl and 2.5 μM RNA. Predictions were made with mFold [16], which predicted only one structure for each RNA. The measured Tm and ΔG° values were calculated based on thermal melting curves (n = 3, +/- standard error of the mean). Values obtained at 180 mM NaCl, at different RNA concentrations, and from assays performed in reverse (transitions from high to low temperature) are shown in S1A Fig. *ND- Not determined due to lack of detectable structure. #The measured values for the 3’-5bp-strong RNA are based on a single inflection point at 66°C, but the melting curve is biphasic (see panel C). Therefore, these values exaggerate the stability of this structure. (C) Representative melting curves at 1 M NaCl and 2.5 μM RNA.