Rapid proton-detected NMR assignment for proteins with fast magic angle spinning.
Rapid proton-detected NMR assignment for proteins with fast magic angle spinning.
J Am Chem Soc. 2014 Aug 7;
Authors: Barbet-Massin E, Pell AJ, Retel J, Andreas LB, Jaudzems K, Franks WT, Nieuwkoop AJ, Hiller M, Higman VA, Guerry P, Bertarello A, Knight MJ, Felletti M, Le Marchand T, Kotelovica S, Akopjana I, Tars K, Stoppini M, Bellotti V, Bolognesi M, Ricagno S, Chou JJ, Griffin RG, Oschkinat H, Lesage A, Emsley L, Herrmann T, Pintacuda G
Abstract
Using a set of six 1H-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 2H/1H exchange, high magnetic fields, high-spinning frequencies (?r/2?>=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 13C/15N 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.
PMID: 25102442 [PubMed - as supplied by publisher]
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