Understanding the binding induced folding of intrinsically disordered proteins by protein engineering: Caveats and pitfalls

Many proteins lack a well-defined three-dimensional structure in isolation. These proteins, typically denoted as intrinsically disordered proteins (IDPs), may display a characteristic disorder-to-order transition when binding their physiological partner(s). From an experimental perspective, it is of great importance to establish the general grounds to understand how such folding processes may be explored. Here we discuss the caveats and the pitfalls arising when applying to IDPs one of the key techniques to characterize the folding of globular proteins, the Phi value analysis. This method is based on measurements of the free energy changes of transition and native states upon conservative, non-disrupting, mutations. On the basis of available data, we reinforce the validity of Phi value analysis in the study of IDPs and suggest future experiments to further validate this powerful experimental method.