Protein resonance assignment at MAS frequencies approaching 100 kHz: a quantitative comparison of J-coupling and dipolar-coupling-based transfer methods

Abstract

We discuss the optimum experimental conditions to obtain assignment spectra for solid proteins at magic-angle spinning (MAS) frequencies around 100 kHz. We present a systematic examination of the MAS dependence of the amide proton T 2′ times and a site-specific comparison of T 2′ at 93 kHz versus 60 kHz MAS frequency. A quantitative analysis of transfer efficiencies of building blocks, as they are used for typical 3D experiments, was performed. To do this, we compared dipolar-coupling and J-coupling based transfer steps. The building blocks were then combined into 3D experiments for sequential resonance assignment, where we evaluated signal-to-noise ratio and information content of the different 3D spectra in order to identify the best assignment strategy. Based on this comparison, six experiments were selected to optimally assign the model protein ubiquitin, solely using spectra acquired at 93 kHz MAS. Within 3 days of instrument time, the required spectra were recorded from which the backbone resonances have been assigned to over 96 %.

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Acknowledgments

We thank Riccardo Cadalbert for expressing the protein and Emilie Testori and Alons Lends for help with crystallization and rotor filling. This work was supported by the Agence Nationale de la Recherche (ANR-11-BSV8-021-01, ANR-12-BS08-0013-01), the ETH Zurich, the Swiss National Science Foundation (Grant 200020_124611), the Centre National de la Recherche Scientifique (CNRS), PUT126 and ETF9229 from Estonian Ministry of Science and Education.

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Correspondence to Ago Samoson or Anja Böckmann or Matthias Ernst or Beat H. Meier.

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Penzel, S., Smith, A.A., Agarwal, V. et al. Protein resonance assignment at MAS frequencies approaching 100 kHz: a quantitative comparison of J-coupling and dipolar-coupling-based transfer methods. J Biomol NMR 63, 165–186 (2015). https://doi.org/10.1007/s10858-015-9975-y

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Keywords

  • Solid-state NMR
  • MAS
  • Assignment