Figure 1. The mutate-and-map method gives an information-rich picture of RNA structure.

a, Mutating a nucleotide and mapping chemical accessibility reveals interactions in the three-dimensional structure of the RNA. The traces are for wild-type (blue) and C18G-mutated (red) variants of the adenine-binding domain of the add riboswitch. These 2′-OH acylation (SHAPE) data were read out by reverse transcription with fluorescently labelled primers and capillary electrophoresis; peaks (left to right) correspond to nucleotides from the 5′ to 3′ end of the RNA. Arrows mark exposure of the mutation site (C18) and of sequence-distant regions brought near this nucleotide by base-pairing (partner G78). b, Entire mutate-and-map data set across 71 single mutations, plotted in grey scale, revealing numerous elements of riboswitch structure. Dark features highlight: (I) the main diagonal stripe showing localized perturbations following C18G mutation; (II–IV) punctate features marking base pairs C18–G78, C26–G44 and C54–G72 in three different helices; (V–VII) more delocalized effects upon helix mutations G14C, G44C and G59C; (VIII) large-scale changes from C69G mutation due to secondary structure rearrangement; (IX) perturbations consistent with loss of adenine binding in A52U variant; (X) evidence for long-range tertiary contact between L2 and L3 upon mutation of C60 and C61 in L3; (XI) ‘symmetric’ mutations in L2 that affect L3; (XII) evidence for U22–A52 base pair in the adenine binding site.