Improved pulse sequences for pure exchange solid-state NMR spectroscopy.
Spin-exchange experiments are useful for improving the resolution and establishment of sequential assignments in solid-state NMR spectra of uniformly (15)N-labeled proteins oriented macroscopically in phospholipid bilayers. To exploit this advantage fully, it is crucial that the diagonal peaks in the two-dimensional exchange spectra are suppressed. This may be accomplished using the recent pure-exchange (PUREX) experiments, which, however, suffer from up to a threefold reduction of the cross-peak intensity relative to experiments without diagonal-peak suppression. This loss in sensitivity may severely hamper the applicability for the study of membrane proteins. In this paper, we present a two-dimensional exchange experiment (iPUREX) which improves the PUREX sensitivity by 50%. The performance of iPUREX is demonstrated experimentally by proton-mediated (15)N-(15)N spin-exchange experiments for a (15)N-labeled N-acetyl-L-valyl-L-leucine dipeptide. The relevance of exchange experiments with diagonal-peak suppression for large, uniformly (15)N-labeled membrane proteins in oriented phospholipid bilayers is demonstrated numerically for the G-protein coupled receptor rhodopsin.
Vosegaard
Thomas
T
Interdisciplinary Nanoscience Center (iNANO) and Laboratory for Biomolecular NMR Spectroscopy, Department of Molecular Biology, University of Aarhus, Aarhus, Denmark. tv@chem.au.dk
Nielsen
Niels C
NC
D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000295:instrumentation / Q000379:methods; D008565:Membrane Proteins / Q000737:chemistry*
D008565:Membrane Proteins