Extended Data Fig. 2: Structure quality of the Tetrahymena ribozyme cryo-EM maps.

(a) Visual inspection of the structure quality of the 2.2 Å map at different contour levels. (b) Four representative nucleotides extracted from the 2.2 Å map with high Q-scores. Each nucleotide is displayed at two density contour levels. The Q-scores for each nucleotide (gray), the backbone (orange), and the base (blue) are shown. The directional orientation of exocyclic nitrogen and oxygen densities pointing out of the bases allowed for differentiation of adenine and guanine. However, distinguishing cytosine and uracil remained difficult because their only difference lies in the exocyclic N4 (cytosine) or O4 (uracil) position. Unambiguously distinguishing between cytosine and uracil may require a resolution closer to 1 Å; for example, the difference between oxygen and nitrogen atoms becomes clear in the 1.34 Å cryo-EM map of the protein apoferritin¹⁷. When visually examining each nucleotide, densities of the backbone were less pronounced than the base. (c) Plots of per-nucleotide Q-score (rolling 10 nt average) separated by part of the nucleotide, with base in blue and the backbone in orange. The backbone was further split into the ribose sugar (green), oxygens attached to the phosphorus (pink), and phosphorus (yellow) represented by dotted lines. Nucleotide numbering, x-axis, is colored by domain matching Fig. 1. The Q-score confirmed quantitatively that resolvability is better for the bases than the backbone across the majority of the 2.2 and 2.3 Å maps. In a previous 3.1 Å map (EMDB-31385, PDB: 7ez0)²⁹, the base and backbone were equally resolved, indicating that increasing the resolution to ~2.3 Å is particularly important to accurately model base orientations and hence increased confidence in modeling water interactions.