Optimized set of two-dimensional experiments for fast sequential assignment, secondary structure determination, and backbone fold validation of 13C/15N-labelled proteins.
NMR experiments are presented which allow backbone resonance assignment, secondary structure identification, and in favorable cases also molecular fold topology determination from a series of two-dimensional 1H-15N HSQC-like spectra. The 1H-15N correlation peaks are frequency shifted by an amount +/- omegaX along the 15N dimension, where omegaX is the Calpha, Cbeta, or Halpha frequency of the same or the preceding residue. Because of the low dimensionality (2D) of the experiments, high-resolution spectra are obtained in a short overall experimental time. The whole series of seven experiments can be performed in typically less than one day. This approach significantly reduces experimental time when compared to the standard 3D-based methods. The here presented methodology is thus especially appealing in the context of high-throughput NMR studies of protein structure, dynamics or molecular interfaces.
Bersch
Beate
B
Institut de Biologie Structurale-Jean-Pierre Ebel, UMR5075 CNRS-CEA-UJF, 41, rue Jules Horowitz, 38027 Grenoble Cedex, France.
Rossy
Emmanuel
E
Covès
Jacques
J
Brutscher
Bernhard
B
D001426:Bacterial Proteins / Q000737:chemistry*; D002244:Carbon / Q000737:chemistry*; D008958:Models, Molecular; D009584:Nitrogen / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular; D010088:Oxidoreductases / Q000737:chemistry*; D017510:Protein Folding; D017433:Protein Structure, Secondary
D001426:Bacterial Proteins; D002244:Carbon; D010088:Oxidoreductases; C021551:mercuric reductase; D009584:Nitrogen