Mapping the Conformational Landscape of Urea-Denatured Ubiquitin Using Residual Dipolar Couplings

摘要

Characterization of the unfolded state is a fundamental prerequisite for understanding protein stability and folding. We have investigated local conformational sampling in urea-denatured ubiquitin at pH 2.5 by measuring an extensive set of residual dipolar couplings (RDCs) under conditions of partial molecular alignment. Seven experimental RDCs per peptide unit, including complementary fixed-geometry and interproton (~1H~N-~1H~N and ~1H~N-~1H~α) couplings, were used to investigate the structural properties of the peptide chain. Amino-acid-specific potentials that simultaneously reproduce all RDCs in the molecule are found to sample more extended conformations than the standard statistical coil description. Analysis of ~3J_(HNHα) scalar couplings measured under the same conditions suggests that neither polyproline II nor extended β-regions dominate this additional sampling of extended conformations. Using this approach we propose a model of the conformational landscape throughout the peptide chain of urea-denatured ubiquitin, providing an essential description for understanding the unfolded state.