期刊
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
卷 136, 期 35, 页码 12489-12497出版社
AMER CHEMICAL SOC
DOI: 10.1021/ja507382j
关键词
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资金
- Agence Nationale de la Recherche (ANR) [10-BLAN-713-01]
- Joint Research Activity and Access to Research Infrastructures
- Marie-Curie ITN in the 7th FP of the EC (EAST-NMR) [228461]
- Marie-Curie ITN in the 7th FP of the EC (BioNMR) [261863]
- Marie-Curie ITN in the 7th FP of the EC (SMBP) [211800]
- CNRS (TGIR-RMN-THC) [FR3050]
- Italian Ministry of Research (FIRB) [RBFR109EOS]
- NIH [EB-001960, EB-002026]
- MRC [MR/K000187/1] Funding Source: UKRI
- Medical Research Council [MR/K000187/1] Funding Source: researchfish
- Direct For Biological Sciences
- Div Of Molecular and Cellular Bioscience [0741914] Funding Source: National Science Foundation
Using a set of six H-1-detected triple-resonance NMR experiments, we establish a method for sequence-specific backbone resonance assignment of magic angle spinning (MAS) nuclear magnetic resonance (NMR) spectra of 5-30 kDa proteins. The approach relies on perdeuteration, amide H-2/H-1 exchange, high magnetic fields, and high-spinning frequencies (omega(r)/2 pi >= 60 kHz) and yields high-quality NMR data, enabling the use of automated analysis. The method is validated with five examples of proteins in different condensed states, including two microcrystalline proteins, a sedimented virus capsid, and two membrane-embedded systems. In comparison to contemporary C-13/N-15-based methods, this approach facilitates and accelerates the MAS NMR assignment process, shortening the spectral acquisition times and enabling the use of unsupervised state-of-the-art computational data analysis protocols originally developed for solution NMR.
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