Protein resonance assignment by solid-state NMR based on 1H-detected 13C-based double-quantum spectroscopy at fast MAS

Abstract

Solid-state NMR spectroscopy is a powerful technique to study insoluble and non-crystalline proteins and protein complexes at atomic resolution. The development of proton (1H) detection at fast magic-angle spinning (MAS) has considerably increased the analytical capabilities of the technique, enabling the acquisition of 1H-detected fingerprint experiments in few hours. Here an approach based on double-quantum (DQ) 13C spectroscopy, detected on 1H, is introduced at fast MAS (70 kHz) to perform the sequential assignment of insoluble proteins of small size, without any specific deuteration requirement. By combining two three-dimensional 1H detected experiments correlating a 13C DQ dimension respectively to its intra-residue and sequential 15N-1H pairs, a sequential walk through DQ (C+CO) resonance is obtained. Our approach takes advantage of fast MAS to achieve an efficient sensitivity and the addition of a DQ dimension provides spectral features useful for the resonance assignment process.