Extended Data Fig. 8: Analysis of overlap solvent shell density.

(a-b) Description of how SWIM-assigned waters (a) from the 2.2 Å model match the 2.3 Å model and map and (b) water from the 2.3 Å model match the 2.2 Å model and map. The top pie chart describes how the waters overlap with waters or ions in the other model after a local superposition, with red indicating overlap, same binding site and superimposable, with water in the other model, orange indicating the same binding site only and brown indicating superimposable only (Methods). Likewise for Mg²⁺ ions in the other model, the same categories are colored green, purple, blue respectively. For the waters with no overlap in the model, the bottom pie chart describes whether the water when placed in the other map has a geometry (purple), density value (> 5σ, green), and/or resolvability (Q-score > 0.70, blue, brown, yellow for full, both half maps, one half map respectively) that pass the SWIM criteria. (c) A specific example of how a Mg²⁺ ion can overlap with a water displayed as in Fig. 5. The cryo-EM densities have the star-shaped shape indicative of a Mg²⁺ ion with a full-coordination shell. The MD agrees that this site contains a Mg²⁺ ion coordinated to the two phosphate and 4 waters. Due to the blurring of density automated SWIM identified two different peak positions within the diffuse density, and modeled a Mg²⁺ ion in the 2.2 Å map and a water in the 2.3 Å map. (d–i) The agreement between the reference map, the 2.2 Å cryo-EM map, and the comparison maps, 2.3 Å cryo-EM map (blue), 2.2 Å model (orange; created by molmap), the MD water density (red), and the 2.2 Å cryo-EM map with the voxel values shuffled (gray). The blue line represents experimental uncertainty while the gray represents performance of a random algorithm. Voxels are defined as “positive” if they have a value above the given contour, 3σ for (d-e), in the reference map and “negative” otherwise. Other metrics are tabulated in Supplementary Table 2. (d-e) The contour of the comparison map is varied to plot (d) Precision-recall (PRC) and (e) receiver operating characteristic (ROC) curves. The Area Under the Curve (AUC) is labeled. (f) The AUC is calculated for a variety of reference map, the 2.2 Å cryo-EM map, contours and for the solvent shell. Trends are consistent over reasonable contours of the reference map, the 2.2 Å map. (g) The model is colored by per-nucleotide experimental precision, defined as the AU-PRC for the independent map. This shows precision decreasing radially from the center of mass of the molecule. (h-i) The local agreement of (h) the SWIM model and (i) the MD density to the reference map, the 2.2 Å cryo-EM map, is plotted, normalized AU-PRC such that a score of 1 is equal to experimental uncertainty (see Methods). The normalized AU-PRC is plotted on the structure with a view of the catalytic core (top) and the tetraloop receptor (bottom), from low agreement (red) to high agreement (blue). Nucleotides with water structure with high experimental uncertainty (AU-PRC^exp < 0.2) are white.