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

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Abstract
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. Copyright © 2006 John Wiley & Sons, Ltd.