Crossing the Entropic Barrier to Coupled Folding and Binding

Intrinsically disordered proteins (IDPs) function in many important cellular pathways, and their disruption contributes to many human diseases. IDPs display minimal if any secondary or tertiary structure in their unbound forms, but will often fold into ordered structures upon binding. It is believed that the coupling of folding and binding is important in balancing the affinity and specificity of the interaction due to the reduction in conformational entropy of the residues at the binding interface. Potential mechanisms include induced folding and conformational selection, which are not mutually exclusive and the predominant method utilized may be biased by the entropic barrier in coupled folding and binding. A model system using the transactivation domain of the tumor suppressor p53 (p53TAD) and its binding partners, the ubiquitin ligases MDM2 and its homologue MDMX, was designed. p53TAD is an IDP that folds when binding with MDM2/X. The conformational entropy of p53TAD is systematically varied using mutagenesis and these mutations result in significant and predictable changes in the binding affinity. This is the first attempt to quantify how differences in the conformational entropy of an IDP affect the binding affinity of a coupled folding reaction, and should give a better understanding of the mechanisms involved in this kind of binding. The importance of this research is highlighted by the high frequency in which folding is coupled to binding in protein-protein interactions.