An early association between the alpha-helix of the TEAD binding domain of YAP and TEAD drives the formation of the YAP:TEAD complex.

The Hippo pathway is an evolutionarily conserved signalling pathway that is involved in the control of organ size and development. The TEAD transcription factors are the most downstream elements of the Hippo pathway, and their transcriptional activity is regulated via the interaction with different co-regulators such as YAP. The structure of the YAP:TEAD complex shows that YAP binds to TEAD via two distinct secondary structure elements, an alpha-helix and an Omega-loop, and site-directed mutagenesis experiments revealed that the Omega-loop is the "hot spot" of this interaction. While substantial knowledge has been gained on how YAP and TEAD interact with each other, little is known about the mechanism leading to the formation of a complex between these two proteins. Here we combine site-directed mutagenesis with pre-steady-state kinetic measurements to show that the association between these proteins follows an apparent one-step binding mechanism. Furthermore, linear free energy relationships and a Phi analysis suggest that binding-induced folding of the YAP alpha-helix to TEAD occurs independently of and before formation of the Omega-loop interface. Thus, the binding-induced folding of YAP appears not to conform to the concomitant formation of tertiary structure (nucleation-condensation) usually observed for coupled binding and folding reactions. Our findings demonstrate how a mechanism reminiscent of the classical framework (diffusion-collision) mechanism of protein folding may operate in disorder-to-order transitions involving intrinsically disordered proteins.