Heat Effects on Coil Hydrodynamic Size Reveal the Energetics of Denatured State Conformational Bias

Abstract

Conformational equilibria in the protein denatured state has key roles regulating folding, stability, and function. Yet detailed information on the denatured state has proven elusive because of the low solubility of unfolded protein and the difficulty in deciphering the character of individual conformations from the ensemble average. To better understand the structural preferences of the denatured state, we have analyzed both the sequence dependence to the mean hydrodynamic size of disordered proteins in water and the impact of heat on the coil dimensions, showing that the sequence dependence and thermodynamic energies associated with intrinsic biases for the α and PPII backbone conformations can be obtained. These results are important for two reasons. First, survey of the protein coil library reveals that the α and PPII basins of the Ramachandran map are the dominant structural features of the protein denatured state. Our results demonstrate how to measure the temperature dependence to the α and PPII populations. Second, it has long been known that the dimensions of denatured proteins in 6 M guanidinium chloride are successfully predicted by the random coil model, irrespective of numerous experiments showing they can maintain residual structures. Hence, the protein hydrodynamic size is thought to give poor structural detail. Our results show that this is not the case for unfolded proteins in water. Rather, surprisingly, the hydrodynamic size of the unfolded state is found to be an effective reporter on the extent of the biases for the α and PPII backbone conformations.