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 H-1-N-15 HSQC-like spectra. The H-1-N-15 correlation peaks are frequency shifted by an amount +/- omega(X) along the N-15 dimension, where omega(X) is the C-alpha, C-beta, or H-alpha 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.