Anisotropic Dynamics of Protein Side Chain NH₂ Groups Revealed through Analysis of ²H and ¹⁵N NMR Relaxation Rates

Although nuclear magnetic resonance (NMR) spectroscopy is powerful for protein dynamics investigations, the anisotropy of internal motions has been difficult to analyze with NMR. In principle, NMR order parameters for multiple bond vectors fixed on the same plane can reveal the anisotropy of internal motions. We investigated the anisotropic dynamics of protein asparagine (Asn) and glutamine (Gln) side chain NH₂ groups using ²H and ¹⁵N NMR relaxation rates. Hindered rotations about the C-N bond causing chemical exchange between the two hydrogens of Asn/Gln NH₂ groups are far slower than ²H relaxation. Using the ²H and ¹⁵N relaxation data at two magnetic fields, we determined two order parameters and ²H quadrupolar coupling constants for each NH₂ group of ubiquitin. Our data clearly illuminate the heterogeneous dynamic properties of the protein side chain NH₂ groups with different degrees of the motional anisotropy, which depends strongly on the local environment.