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Selectively dispersed isotope labeling for protein structure determination by magic angle spinning NMR

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Abstract

The power of nuclear magnetic resonance spectroscopy derives from its site-specific access to chemical, structural and dynamic information. However, the corresponding multiplicity of interactions can be difficult to tease apart. Complimentary approaches involve spectral editing on the one hand and selective isotope substitution on the other. Here we present a new “redox” approach to the latter: acetate is chosen as the sole carbon source for the extreme oxidation numbers of its two carbons. Consistent with conventional anabolic pathways for the amino acids, [1-13C] acetate does not label α carbons, labels other aliphatic carbons and the aromatic carbons very selectively, and labels the carboxyl carbons heavily. The benefits of this labeling scheme are exemplified by magic angle spinning spectra of microcrystalline immunoglobulin binding protein G (GB1): the elimination of most J-couplings and one- and two-bond dipolar couplings provides narrow signals and long-range, intra- and inter-residue, recoupling essential for distance constraints. Inverse redox labeling, from [2-13C] acetate, is also expected to be useful: although it retains one-bond couplings in the sidechains, the removal of CA–CO coupling in the backbone should improve the resolution of NCACX spectra.

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Acknowledgments

We thank Dr. Alexei Belenky for assistance with LC–MS, Dr. Christopher Turner for assistance with the COSY and HSQC experiments, Loren Andreas, Dr. Marcel Reese, and Dr. Yongchao Su for assistance with the set up of the 800 MHz spectrometer, and Dr. Jochem Struppe for assistance with the TEDOR pulse program. Research reported in this publication was supported by the National Institute of Biomedical Imaging and Bioengineering of the National Institutes of Health under award numbers EB001035, EB001960 and EB002026. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

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Authors and Affiliations

  1. Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA, USA

    Matthew T. Eddy & Robert G. Griffin

  2. Francis Bitter Magnet Laboratory, Massachusetts Institute of Technology, Cambridge, MA, USA

    Matthew T. Eddy, Astrid C. Sivertsen & Robert G. Griffin

  3. Department of Chemistry, Brandeis University, Waltham, MA, USA

    Marina Belenky, Astrid C. Sivertsen & Judith Herzfeld

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  1. Matthew T. Eddy
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  2. Marina Belenky
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  3. Astrid C. Sivertsen
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  4. Robert G. Griffin
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  5. Judith Herzfeld
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Correspondence to Judith Herzfeld.

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Eddy, M.T., Belenky, M., Sivertsen, A.C. et al. Selectively dispersed isotope labeling for protein structure determination by magic angle spinning NMR. J Biomol NMR 57, 129–139 (2013). https://doi.org/10.1007/s10858-013-9773-3

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