Direct Measurement of 15N Chemical Shift Anisotropy in Solution

The magnitude and orientation of the 15 N chemical shift anisotropy (CSA) tensors are determined for human ubiquitin in solution from 15 N 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/ R 2, of the cross correlation (η) between 15 N CSA and 15 N− 1 H dipolar interaction and of the rate ( R 2 ) of 15 N transverse relaxation. Since the η/ R 2 ratio does not contain any direct dependence on protein dynamics, the present approach is free from assumptions about overall and local motions. The 15 N 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 15 N CSA tensor was 15.7 ± 5.0° (range 6−26°). The results indicate the importance of residue-specific 15 N CSA for accurate analysis of dynamics from relaxation data, and provide access to the CSA in solution, which may be structurally useful.