13C NMR studies of the molecular dynamics of selectively 13C-enriched ribonuclease complexes

13C spin-lattice (T1) relaxation times determined at four frequencies (25, 68, 100, and 125 MHz) have been used to probe the molecular dynamics of ribonuclease S' complexes prepared from synthetic amino-terminal peptides containing 13C enrichment (ca. 90%) at selected sites [Niu, C., Matsuura, S., Shindo, H., & Cohen, J. S. (1979) J. Biol. Chem. 254, 3788]. It was found that the motion of the C alpha-H bond of Ala-5 could not be determined by isotropic reorientation alone. The time scale and spatial restriction on the internal motion of this residue were determined by the model-free approach of Lipari and Sazbo [Lipari, G., & Szabo, A. (1982) J. Am. Chem. Soc. 104, 4546-4559]. It was found that the C alpha-H bond, in addition to an overall correlation time of 20 ns, underwent internal motion with a correlation time of 0.5 ns and a generalized order parameter S corresponding to a cone semiangle of 23 degrees C. The C beta-H bond had a correlation time of 37 ps, reflecting the fast rotation of the methyl group, and had an S value close to that expected if the C alpha-C beta and C alpha-H bonds have the same degree of spatial restriction.