MOLECULAR RECOGNITION MECHANISM OF p63 BY ITCH-E3 LIGASE: ADVANCES AND EFFECTS OF A p63 MUTATION RELATED TO ECTODERMAL DYSPLASIAS.

Recently, it has been shown that Itch mediates the degradation of TAp63 and ΔNp63 proteins1. Itch E3–ligase contains four WW domains important in the recognized process. Several signalling complexes, that these domains mediate, have been implicated in human diseases (Muscular Dystrophy, Alzheimer's Disease, Huntington Disease etc.). WW domains are highly compact protein-protein binding modules that interact with short proline-rich sequences. Based on their ligand-binding specificity they have been categorized into four groups. WW domains fold into stable three-stranded antiparallel b-sheet structures, and their primary sequence share two conserved tryptophan residues spaced 20-22 amino acids apart. The four WW domains of Itch are considered belonging to the Group I, which binds polypeptides with a PY motif characterized by a PPXY consensus sequence, where X can be any residue. It is likely that the Itch-p63 interaction results from a direct interaction of Itch-WW2 domain with the PY motif of p63. Here, we present a structural characterization of the interaction by fluorescence, CD and NMR spectroscopy of the Itch-WW2 domain. Interaction studies in vitro between Itch-WW2 domain and pep63, which correspond to the fragment of the p63 protein including the PY motif, were performed. Moreover, the effects of a site specific mutation of p63, that has been reported in both Hay–Wells syndrome and Rapp–Hodgkin syndrome, was also evaluated both on the conformation of pep63 and on the WW-pep63 interaction. 1Rossi M., Aqeilan I., Neale M., Candi E., Salomoni P., Knight R.A., Croce C.M., Melino G. PNAS (2006) 103: 12753-58