Abstract
Magic-angle spinning (MAS) solid-state NMR (SSNMR) spectroscopy of uniformly-13C,15N labeled protein samples provides insight into atomic-resolution chemistry and structure. Data collection efficiency has advanced remarkably in the last decade; however, the study of larger proteins is still challenged by relatively low resolution in comparison to solution NMR. In this study, we present a systematic analysis of SSNMR protein spectra acquired at 11.7, 17.6 and 21.1 Tesla (1H frequencies of 500, 750, and 900 MHz). For two protein systems—GB1, a 6 kDa nanocrystalline protein and DsbA, a 21 kDa nanocrystalline protein—line narrowing is demonstrated in all spectral regions with increasing field. Resolution enhancement is greatest in the aliphatic region, including methine, methylene and methyl sites. The resolution for GB1 increases markedly as a function of field, and for DsbA, resolution in the C–C region increases by 42%, according to the number of peaks that can be uniquely picked and integrated in the 900 MHz spectra when compared to the 500 MHz spectra. Additionally, chemical exchange is uniquely observed in the highest field spectra for at least two isoleucine Cδ1 sites in DsbA. These results further illustrate the benefits of high-field MAS SSNMR spectroscopy for protein structural studies.
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Acknowledgments
The authors thank the National Institute of Heath for funding through NIGMS (GM073770), NIGMS/Roadmap Initiative (GM075937) and Molecular Biophysics Training Grant (to LJS and AJN), David Hoyt, Jesse Sears, and Paul Ellis at the Environmental Molecular Science Laboratory (a national scientific user facility sponsored by the Department of Energy Office of Biological and Environmental Research) located at Pacific Northwest National Laboratory and operated for DOE by Batelle for their assistance in acquiring the 900 MHz data, Dr. Donghua Zhou for pulse sequence code, Dr. Trent Franks and Benjamin Fisher of the VOICE NMR Facility for technical assistance.
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Sperling, L.J., Nieuwkoop, A.J., Lipton, A.S. et al. High resolution NMR spectroscopy of nanocrystalline proteins at ultra-high magnetic field. J Biomol NMR 46, 149–155 (2010). https://doi.org/10.1007/s10858-009-9389-9
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DOI: https://doi.org/10.1007/s10858-009-9389-9