Yorkie-Warts Complexes are an Ensemble of Interconverting Conformers Formed by Multivalent Interactions

Abstract

WW domain and PPXY motif sequences are often presented in multiple copies in several proteins at the critical homeostasis juncture of cell death versus cell replication. While biophysical studies of single WW domain-single PPXY motif complexes abound in the literature, the molecular mechanisms of multivalent WW domain-PPXY assemblies are still poorly understood. by way of investigating such assemblies, we have characterized the multivalent association of the entire cognate binding domains, two WW domains and five PPXY motifs respectively, of the Yorkie transcription coactivator and the Warts tumor suppressor. Isothermal titration calorimetry and native-state mass spectrometry of Yorkie interactions with the full-length Warts PPXY domain, and numerous PPXY motif variants of Warts show that the two proteins assemble via binding of the tandem WW domains to adjacent PPXY pairs to produce an ensemble of interconverting complexes of variable stoichiometries, binding energetics and WW domain occupancy. Apparently, the Yorkie tandem WW domains first target the two adjacent PPXY motifs at the C- terminus of the Warts polypeptide and additional WW domains bind unoccupied motifs. The ensemble of interconverting conformers may be common in multivalent WW domain-PPXYinteractions, and may promote the adaptability and versatility of WW domain-PPXY mediated cellular processes.