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J Am Chem Soc. 2010 Feb 24;132(7):2138-9. doi: 10.1021/ja910523q.

Diagonal-free 3D/4D HN,HN-TROSY-NOESY-TROSY.

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NMR platform, CiC bioGUNE, Parque Tecnologico de Bizkaia, Ed. 800, 48160 Derio, Spain.


Structural biology by NMR spectroscopy relies on measuring interproton distances via NOE cross-signals in nuclear Overhauser effect spectroscopy (NOESY) spectra. In proteins, the subset of H(N)-H'(N) NOE contacts is most important for deriving initial structural models and for spectral assignment by "NOE walking". Here we present a fully optimized NMR experiment for measuring these pivotal contacts: diagonal-free 3D/4D HN,HN-TROSY-NOESY-TROSY. It combines all of the critical requirements for extracting the optimal H(N)-H'(N) distance information: the highest resolution by consistent transverse relaxation-optimized spectroscopy (TROSY) evolution, the largest spectral dispersion in two (15)N dimensions, and maximal coverage and purity through specific suppression of the intense diagonal signals that are the main source of overlap, artifacts, and bias in any NOESY spectrum. Most notably, diagonal suppression here comes without compromising the NOE cross-signal intensities. This optimized experiment appears to be ideal for a broad range of structural studies, particularly on large deuterated, partially unfolded, helical, and membrane proteins.

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