Imino hydrogen positions in nucleic acids from density functional theory validated by NMR residual dipolar couplings

Hydrogen atom positions of nucleotide bases in RNA structures solved by X-ray crystallography are commonly derived from heavy-atom coordinates by assuming idealized geometries. In particular, N1-H1 vectors in G and N3-H3 vectors in U are commonly positioned to coincide with the bisectors of their respective heavy-atom angles. We demonstrate that quantum-mechanical optimization of the hydrogen positions relative to their heavy-atom frames considerably improves the fit of experimental residual dipolar couplings to structural coordinates. The calculations indicate that deviations of the imino N-H vectors in RNA U and G bases result from H-bonding within the base pair and are dominated by the attractive interaction between the H atom and the electron density surrounding the H-bond-acceptor atom. DFT optimization of H atom positions is impractical in structural biology studies. We therefore have developed an empirical relation that predicts imino N-H vector orientations from the heavy-atom coordinates of the base pair. This relation agrees very closely with the DFT results, permitting its routine application in structural studies.