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Estimating side-chain order in methyl-protonated, perdeuterated proteins via multiple-quantum relaxation violated coherence transfer NMR spectroscopy


Abstract Relaxation violated coherence transfer NMR spectroscopy (Tugarinov et al. in J Am Chem Soc 129:1743â??1750, 2007) is an established experimental tool for quantitative estimation of the amplitudes of side-chain motions in methyl-protonated, highly deuterated proteins. Relaxation violated coherence transfer experiments monitor the build-up of methyl proton multiple-quantum coherences that can be created in magnetically equivalent spin-systems as long as their transverse magnetization components relax with substantially different rates. The rate of this build-up is a reporter of the methyl-bearing side-chain mobility. Although the build-up of multiple-quantum 1H coherences is monitored in these experiments, the decay of the methyl signal during relaxation delays occurs when methyl proton magnetization is in a single-quantum state. We describe a relaxation violated coherence transfer approach where the relaxation of multiple-quantum 1Hâ??13C methyl coherences during the relaxation delay period is quantified. The NMR experiment and the associated fitting procedure that models the time-dependence of the signal build-up, are applicable to the characterization of side-chain order in [13CH3]-methyl-labeled, highly deuterated protein systems up to ~100 kDa in molecular weight. The feasibility of extracting reliable measures of side-chain order is experimentally verified on methyl-protonated, perdeuterated samples of an 8.5-kDa ubiquitin at 10°C and an 82-kDa Malate Synthase G at 37°C.

• Content Type Journal Article
• Category Article
• Pages 1-11
• DOI 10.1007/s10858-012-9604-y
• Authors
  • Hechao Sun, Department of Chemistry and Biochemistry, Center for Biomolecular Structure and Organization, University of Maryland, Biomolecular Sci. Bldg., College Park, MD 20742, USA
  • Raquel Godoy-Ruiz, Department of Chemistry and Biochemistry, Center for Biomolecular Structure and Organization, University of Maryland, Biomolecular Sci. Bldg., College Park, MD 20742, USA
  • Vitali Tugarinov, Department of Chemistry and Biochemistry, Center for Biomolecular Structure and Organization, University of Maryland, Biomolecular Sci. Bldg., College Park, MD 20742, USA

• • Journal Journal of Biomolecular NMR
  • Online ISSN 1573-5001
  • Print ISSN 0925-2738

Source: Journal of Biomolecular NMR