Solution NMR Measurement of Amide Proton Chemical Shift Anisotropy in 15N-Enriched Proteins. Correlation with Hydrogen Bond Length

Cross-correlation between 15 N− 1 H dipolar interactions and 1 H N chemical shift anisotropy (CSA) gives rise to different relaxation rates of the doublet components of 1 H−{ 15 N} peptide backbone amides. Two schemes for quantitative measurement of this effect are described and demonstrated for samples of uniformly 15 N-enriched ubiquitin and perdeuterated 15 N-enriched HIV-1 protease. The degree of relaxation interference correlates with the isotropic 1 H N chemical shift, and results indicate that an upfield change of the most shielded principal components of the CSA tensor is correlated with an approximately 2-fold larger downfield shift of the average of the other two components. The magnitude of the relaxation interference is large in β-sheet and considerably smaller in α-helices. This correlation is not dominated by the backbone geometry but reflects the slightly longer hydrogen bond length in helices compared to β-sheet. The smallest relaxation interference effect in ubiquitin is observed for Ser 20 -H N and Ile 36 -H N, which are the only two amide protons that are not hydrogen bonded in the crystal structure of ubiquitin, inaccessible to solvent, and not highly mobile.