Proteins adopt non-native geometries during their synthesis, folding, transport, and turnover and a significant fraction of the proteome is intrinsically disordered. The dimensions that unfolded and intrinsically disordered proteins (IDPs) adopt at low or no denaturant has been controversial. We recently developed an analysis procedure for small-angle X-ray scattering (SAXS) and found that even relatively hydrophobic IDPs remain nearly as expanded as the chemically denatured ensemble. Hence, water should be considered to be a good solvent for unfolded states of foldable sequences. While advances in SAXS and FRET analysis has narrowed the difference in their views of the dimensions of unfolded states, here we show that fluorophores used in FRET studies contribute to some of the discrepancy. We also tested the suggestion that FRET and SAXS results may be reconciled if the Rg can vary independently from the end-to-end distance (Ree). Our analysis finds that these two parameters are coupled and SAXS can accurately extract Rg, solvent quality (Flory exponent nu) and Ree even for heteropolymeric sequences in a single measurement. We conclude that mild chain contraction and fluorophore-based interactions at lower denaturant concentrations, along with improved analysis procedures for both SAXS and FRET, can explain the preponderance of existing data regarding the dimensions of unfolded polypeptides.

Using SAXS to measure the solvent quality of water for disordered proteins as well as the collapse induced by FRET fluorophores

2019 Spring National Meeting

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