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Measurement of two- and three-bond 13C−1H J couplings to the Cδ carbons of leucine residues in staphylococcal nuclease

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Summary

A new 1H-detected 3D NMR experiment is described that permits quantitative measurement of two- and three-bond 13C−1H couplings in proteins with selectively 13C-enriched methyl sites. The method is demonstrated for staphylococcal nuclease selectively [5,5 13C]-labeled in all 11 leucine positions and ligated with thymidine 3′,5′-biphosphate and Ca2+. Two- and three-bond 13C methyl-proton couplings are reported and, together with the measured three-bond JCαCδ in uniformly 13C-enriched staphylococcal nuclease, the χ 2- and the stereospecific assignments of the Cδ methyl group with respect to the prochiral β-protons were determined. The same residues that were previously found to have high degrees of internal mobility on the basis of 13C relaxation times have measured coupling constants that are indicative of motional averaging.

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Authors and Affiliations

  1. Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 20892, Bethesda, MD, USA

    Geerten W. Vuister & Ad Bax

  2. Bone Research Branch, National Institute of Dental Research, National Institutes of Health, 20892, Bethesda, MD, USA

    Toshimasa Yamazaki & Dennis A. Torchia

Authors
  1. Geerten W. Vuister
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  2. Toshimasa Yamazaki
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  3. Dennis A. Torchia
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  4. Ad Bax
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Vuister, G.W., Yamazaki, T., Torchia, D.A. et al. Measurement of two- and three-bond 13C−1H J couplings to the Cδ carbons of leucine residues in staphylococcal nuclease. J Biomol NMR 3, 297–306 (1993). https://doi.org/10.1007/BF00212516

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  • DOI: https://doi.org/10.1007/BF00212516

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