Figure 1.

Definition of the reference Cartesian frame on the nucleobases and of the parameters used to define the position of the O^W relatively to the nucleobase. The origin is at the geometrical center of the heterocycle skeleton, the x-axis passing through the N3 atom for pyrimidines and through the middle point of N1–C2 bond for purines, the y-axis forms a 90° angle with the x-axis, with the C6 atom of purines and the C4 atom of pyrimidines lying in the xy-plane at positive y values, the z-axis is the cross product of the versors (unit vectors indicating the directions) along the x and y-axes, thus forming a right-handed frame. The orange curve defines a circle of radius 1.5 Å in the xy-plane of pyrimidines, and an ellipse with minor and major axes equal to 1.5 and 2.5 Å in the xy-plane of purines. A water and a nucleobase are considered to be interacting if the projection of O^W on the xy-plane is within the orange circle/ellipse, with the vertical distance (along the z axis) in the −4.0 to +4.0 Å range. The instances of O^W-nucleobase stacking contacts with specification of their PDB IDs and residues involved are reported in Supplementary Table S1.