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

Abstract

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.

Global and internal molecular dynamics of poly(acrylamide-co-allyl 2-acetamido-2-deoxy-D-glucopyranoside) glycopolymers from 13C NMR relaxation studies

“…Since analyses have proven useful for the analysis of relaxation data in recent years, a certain preference has been given to the utilization of the "modelfree" approach of Lipari and Szabo (16) (equivalent in spectral density form to the "two-step" model suggested by Wennerstrom et al (17)). A number of authors have applied this model-free approach to the molecular dynamics of a number of biologically related systems (18)(19)(20)(21). In particular, and in relation to this work, the model has been adapted to examine both the internal and overall motions of a number of saccharides (22)(23)(24).…”

Deuterium labelling in NMR structural analysis of larger proteins

“…Nevertheless, recent advances in peptide synthesis (DeGrado & Kaiser, 1980;Aimoto et al 1989) may render this approach worthwhile in selected cases. Various site selective labelling experiments have been carried out making use of limited enzymatic digestion and reconstitution or resynthesis of peptide linkages (Richarz et al 1980;Hughes et al 1984;Fisher et al 1989). However, such an approach does not offer the generality needed for extensive NMR analysis.…”

Tryptophan sidechain dynamics in hydrophobic oligopeptides determined by use of 13C nuclear magnetic resonance spectroscopy

“…NMR relaxation techniques can be used to detect and quantify picosecond motions but are often of limited value due to the low natural abundance of the isotope which necessitates the use of high and frequently unattainable protein concentrations. With selective enrichment the method becomes particularly appealing and measurement of NMR relaxation parameters has clearly demonstrated the occurence of very rapid motions in proteins and peptides (Niu et al, 1979;Eakin et al, 1975;Jones et al, 1976;Blakeley et al, 1978;Deber et al, 1978;Cohen et al, 1979;Harina et al, 1980;Matta et al, 1980;Wooten et al, 1981;Hughes et al, 1984).…”

“…A conceptually simpler, model-free approach was originally applied by Lipari and Szabo (1982a,b) to a few resolvable sidechain resonances of myoglobin, dihydrofolate reductase, and bovine pancreatic trypsin inhibitor. The approach was also used by Hughes et al (1984) in the analysis of '3C spin-lattice relaxation times for ribonuclease S' complexes using selectively '3C-enriched S-peptide. More recently, Henry et al (1986) have applied the methodology to the analysis of backbone dynamics of detergent-solubilized M1 3 coat protein.…”