Influence of Experimental Uncertainties on the Properties of Ensembles Derived from NMR Residual Dipolar Couplings

Ensemble simulations restrained by residual dipolar couplings (RDCs) measured by NMR have in recent years emerged as a powerful strategy for the experimental characterization of the structural heterogeneity of macromolecules. Given the wide range of potential applications of this approach, it is important to determine whether the resulting ensembles are affected by unavoidable experimental uncertainties in the measurement of RDCs. Using the well-characterized protein ubiquitin as a model system, we have assessed the effect of experimental error and conclude that the structural heterogeneity in solution can be accurately characterized when experimental uncertainties in the NH RDCs are lower than 0.5 Hz. Our results indicate that determining native ensembles using ensemble simulations restrained by this NMR parameter is a robust procedure.