Methyl motions in 13C-methylated concanavalin as studied by carbon-13 magnetic resonance relaxation techniques

Abstract

The carbon- 13 spin-lattice relaxation times and nuclear Overhauser enhancements of the N epsilon-monomethyllysine, N epsilon,N epsilon-dimethyllysine, and N alpha,N alpha-dimethylalanine resonances of 13C-methylated concanavalin A have been measured at three carbon frequencies and compared to the relaxation parameters predicted by several motional models. The experimental parameters cannot be reproduced by a simple dipolar relaxation model which includes isotropic reorientation of the protein plus free internal rotational diffusion of the methyl groups but are well predicted by a wobble in a cone model which includes isotropic reorientation of the protein at 33 ns, free internal rotational diffusion of the methyl groups, and a wobble diffusion which reflects the net motion of the amino acid side chains. The analysis indicates that the methylated epsilon-amino side chains exhibit only slightly more motional freedom than does the methylated N-terminal alpha-amino group and suggests some restriction of methyl group rotation in the dimethylamino residues.

NMR studies of the methionine methyl groups in calmodulin

“…The resonances of the Met methyl groups were studied in two-dimensional proton-detected heteronuclear NMR experiments and assigned to specific Met residues by site.dire~1:d mutagenesis (Met~Leu). It has been known for some time that natural abundance and isotope-labelled ~3C relaxation measurements can provide a unique insight into the motions of methyl groups in amino acid sidechains in proteins [16][17][18][19], (for more general reviews see [20,21]). In addition, carbon-13 relaxation lacks some of the disadvantages inherent in proton NMR relaxation studies [22].…”

A Novel Model-Free Analysis of13C NMR Relaxation of Alanine-Methyl Side-Chain Motions in Peptides

“…C NMR relaxation measurements (proton correlation times, restrictions, and correlations between varicoupled and decoupled) were performed at two NMR frequencies ous rotations (12)(13)(14)(15)(16)(17)(18)(19)(20). Methyl-group rotations are sensitive (150 and 62.5 MHz) and over a wide range of temperatures (5 to to protein/peptide conformation in terms of sequentially and 75ЊC) to study motional dynamics of the alanine side-chain methyl spatially neighboring residues (19,20).…”

Angular variances for internal bond rotations of side chains in GXG-based tripeptides derived from13C-NMR relaxation measurements: Implications to protein folding

“…where the correlation time 1 and angular variance 1 were previously defined. For small peptides, the assumption that side-chain bond rotations occur as low amplitude rotational fluctuations occurring within a potential well may not be valid, and other motional models need to be considered for a more realistic picture of the dynamics.…”

“…Rotations about specific side-chain bonds, for example, are often more sensitive to changes in protein conformation and folding than are backbone bond rotations. [1][2][3][4][5][6][7][8][9][10][11][12] In folded proteins and peptides, backbone bond rotations that occur primarily via rotational fluctuations within potential wells are often easier to model, whereas side chains can generally probe greater conformational space and also undergo jumps among various rotomer states. From studies of the temperature dependence of nmrderived motional parameters, it is apparent that in partially folded peptides, 11,12 amplitudes of internal rotations about backbone , bonds vary little with temperature, whereas in folded structures, correlations between various i (t) and k (t) bond rotations can be substantial and can change dramatically with temperature.…”