PMID:12465972
| Citation |
Sibille, N, Bersch, B, Covès, J, Blackledge, M and Brutscher, B (2002) Side chain orientation from methyl 1H-1H residual dipolar couplings measured in highly deuterated proteins. J. Am. Chem. Soc. 124:14616-25 |
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| Abstract |
High-level deuteration is a prerequisite for the study of high molecular weight systems using liquid-state NMR. Here, we present new experiments for the measurement of proton-proton dipolar couplings in CH(2)D methyl groups of (13)C labeled, highly deuterated (70-80%) proteins. (1)H-(1)H residual dipolar couplings (RDCs) have been measured in two alignment media for 57 out of 70 possible methyl containing residues in the 167-residue flavodoxin-like domain of the E. coli sulfite reductase. These data yield information on the orientation of the methyl symmetry axis with respect to the molecular alignment frame. The alignment tensor characteristics were obtained very accurately from a set of backbone RDCs measured on the same protein sample. To demonstrate that accurate structural information is obtained from these data, the measured methyl RDCs for Valine residues are analyzed in terms of chi(1) torsion angles and stereospecific assignment of the prochiral methyl groups. On the basis of the previously determined backbone solution structure of this protein, the methyl RDC data proved sufficient to determine the chi(1) torsion angles in seven out of nine valines, assuming a single-rotamer model. Methyl RDCs are complementary to other NMR data, for example, methyl-methyl NOE, to determine side chain conformation in high molecular weight systems. |
| Links | |
| Keywords |
Carbon Isotopes; Deuterium/chemistry; Escherichia coli/enzymology; Flavodoxin/chemistry; Models, Molecular; Nitrogen Isotopes; Nuclear Magnetic Resonance, Biomolecular/methods; Oxidoreductases Acting on Sulfur Group Donors/chemistry; Protein Conformation; Protein Structure, Tertiary |
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