An approach to direct determination of protein dynamics from ¹⁵N NMR relaxation at multiple fields, independent of variable ¹⁵N chemical shift anisotropy and chemical exchange contributions

An approach to protein dynamics analysis from ¹⁵N relaxation data is demonstrated, based on multiple-field relaxation data. This provides a direct, residue-specific determination of both the spectral density components, the ¹⁵N chemical shift anisotropy (CSA) and the conformational exchange contribution to the ¹⁵N line width. Measurements of R₁, R₂, and ¹⁵N{¹H} NOE are used. The approach is free from any assumption about the values of the CSA or of the conformational exchange. Using this approach, the spectral densities, the values of ¹⁵N CSA, and the conformational exchange contribution to the ¹⁵N line width are directly determined from the relaxation data for human ubiquitin, collected at 360, 500, and 600 MHz. The spectral densities are analyzed in terms of the order parameter and the correlation time of local motion, using an axially symmetric overall rotational diffusion model. The residue-specific values of ¹⁵N CSA and the spectral densities obtained using this approach are in agreement with those derived previously [Fushman, Tjandra, and Cowburn. J. Am. Chem. Soc. 1998, 120, 10947-10952] from CSA/dipolar cross-correlation analysis. Accurate determination of spectral densities and order parameters from ¹⁵N relaxation may be accomplished by analysis of multiple-field data without assumption of constant CSA or zero chemical exchange contributions.