| Abstract
| - Side-chain dynamics in proteins can be characterized by the NMR measurement of 13C and 2Hrelaxation rates. Evaluation of the corresponding spectral densities limits the slowest motions that can bestudied quantitatively to the time scale on which the overall molecular tumbling takes place. A differentmeasure for the degree of side-chain order about the Cα−Cβ bond (χ1 angle) can be derived from 3JC‘-Cγand 3JN-Cγ couplings. These couplings can be measured at high accuracy, in particular for Thr, Ile, and Valresidues. In conjunction with the known backbone structures of ubiquitin and the third IgG-binding domainof protein G, and an extensive set of 13C−1H side-chain dipolar coupling measurements in oriented media,these 3J couplings were used to parametrize empirical Karplus relationships for 3JC‘-Cγ and 3JN-Cγ. TheseKarplus curves agree well with results from DFT calculations, including an unusual phase shift, which causesthe maximum 3JCC and 3JCN couplings to occur for dihedral angles slightly smaller than 180°, particularlynoticeable in Thr residues. The new Karplus curves permit determination of rotamer populations for the χ1torsion angles. Similar rotamer populations can be derived from side-chain dipolar couplings. Conversionof these rotamer populations into generalized order parameters, SJ2 and SD2, provides a view of side-chaindynamics that is complementary to that obtained from 13C and 2H relaxation. On average, results agreewell with literature values for 2H-relaxation-derived Srel2 values in ubiquitin and HIV protease, but also identifya fraction of residues for which SJ,D2< Srel2. This indicates that some of the rotameric averaging occurs ona time scale too slow to be observable in traditional relaxation measurements.
|
