Direct measurement of ¹⁵N chemical shift anisotropy in solution

The magnitude and orientation of the ¹⁵N chemical shift anisotropy (CSA) tensors are determined for human ubiquitin in solution from ¹⁵N relaxation data at 600, 500, and 360 MHz. The analysis uses the model- independent approach [Fushman, D.; Cowburn, D. J. Am. Chem. Soc. 1998, 120, 7109-10] based on a ratio, η/R₂, of the cross correlation (η) between ¹⁵N CSA and ¹⁵N-¹H dipolar interaction and of the rate (R₂) of ¹⁵N transverse relaxation. Since the η/R₂ ratio does not contain any direct dependence on protein dynamics, the present approach is free from assumptions about overall and local motions. The ¹⁵N CSA values fall in the range -125 to -216 ppm, with the average value of -157 ± 19 ppm; the average angle between the NH bond and the unique principal axis of the ¹⁵N CSA tensor was 15.7 ± 5.0°(range 6-26°). The results indicate the importance of residue- specific ¹⁵N CSA for accurate analysis of dynamics from relaxation data, and provide access to the CSA in solution, which may be structurally useful.