Many intrinsically disordered proteins undergo a disorder-to-order transition upon binding to their target molecules. Despite their plasticity, disordered proteins have often been observed in the unbound state to possess latent structure resembling the folded bound state. However, the role of this preformed structure and its effect on the thermodynamics of binding are currently not well understood. Our group has recently shown through temperature-series isothermal titration calorimetry that the folding-upon-binding of a model disordered protein, FCP1, to its target, RAP74, is equivalent to the cooperative folding of a globular protein. Changes in molar enthalpy upon formation of the FCP1–RAP74 complex were deconstructed into contributions from the primary driving force of that interaction, the hydrophobic effect, and other concomitant driving forces, such as those associated with helix formation. In the current work, we attempt to parse the contribution from helix formation through the generation of a helicity series by rational mutagenesis of the disordered protein. In this way, a panel of FCP1 mutants is prepared with amino acid substitutions that confer modulated helicity without disrupting the hydrophobic binding face. Numerous biophysical techniques allow us to use this helicity series to better understand the contribution of latent secondary structure in an intrinsically disordered protein to the thermodynamics of binding to a target molecule.

Examination of latent structure in an intrinsically disordered protein and the thermodynamics of its binding to a target molecule

249th National Meeting (2015)

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