Fig. 8. Secondary structure analysis of the ribozyme GTR1.

(A) SHAPE reactivity profile of the GTR1 ribozyme under optimal reaction conditions. The SHAPE reactivity was determined by reverse transcription of two radiolabeled primers with the SHAPE-reacted RNA. Cutoffs of 0.2 and 0.5 (light blue lines) were chosen to distinguish positions with low reactivity (black), medium reactivity (orange), and high reactivity (red). Error bars are SDs from three independent probing experiments. (B) Secondary structure resulting from calculation with RNAfold in the ViennaRNA package 2.0 (38), using the constraints of the SHAPE reactivity. Color coding is as in (A). The black bar at positions 138 to 141 denote a SHAPE-protected region that may pair with positions 47 to 49 or 86 to 89 (gray bars). Positions labeled in gray did not have SHAPE signals because a signal corresponding to position 1 was too close to full-length extension and because positions 176 to 195 were covered by the reverse transcription primer.