Determination of the Glycosidic Bond Angle χ in RNA from Cross-Correlated Relaxation of CH Dipolar Coupling and N Chemical Shift Anisotropy

A new heteronuclear NMR pulse sequence, the quantitative Γ(HCN) experiment, for the determination of the glycosidic torsion angle χ in ¹³C,¹⁵N-labeled oligonucleotides is described. The Γ(HCN) experiment allows measurement of CH dipole−dipole, N chemical shift anisotropy cross-correlated relaxation rates ( and for pyrimidines and and for purines). A nucleotide-specific parametrization for the dependence of these Γ-rates on χ based on ¹⁵N chemical shift tensors determined by solid-state NMR experiments on mononucleosides (Stueber, D.; Grant, D. M. J. Am. Chem. Soc. 2002, 124, 10539−10551) is presented. For a 14-mer and a 30-mer RNA of known structures, it is found that the Γ(HCN) experiment offers a very sensitive parameter for changes in the angle χ and allows restraining of χ with an accuracy of around 10 degrees for residues which do not undergo conformational averaging. Therefore, the Γ(HCN) experiment can be used for the determination of χ in addition to data derived from ³J(C,H)-coupling constants. As shown for the 30-mer RNA, the derived torsion angle information can be incorporated as additional restraint, improving RNA structure calculations.