HET-SOFAST NMR for fast detection of structural compactness and heterogeneity along polypeptide chains.

Structure elucidation of proteins by either NMR or X-ray crystallography often requires the screening of a large number of samples for promising protein constructs and optimum solution conditions. For large-scale screening of protein samples in solution, robust methods are needed that allow a rapid assessment of the folding of a polypeptide under diverse sample conditions. Here we present HET-SOFAST NMR, a highly sensitive new method for semi-quantitative characterization of the structural compactness and heterogeneity of polypeptide chains in solution. On the basis of one-dimensional 1H HET-SOFAST NMR data, obtained on well-folded, molten globular, partially- and completely unfolded proteins, we define empirical thresholds that can be used as quantitative benchmarks for protein compactness. For 15N-enriched protein samples, two-dimensional 1H-15N HET-SOFAST correlation spectra provide site-specific information about the structural heterogeneity along the polypeptide chain.