Dipartimento di Scienze e Tecnologie Chimiche, University of Rome Tor Vergata, Italy.
Cell Cycle. 2010 Sep 15;9(18):3730-9. Epub 2010 Sep 8.
The E3 ubiquitin ligase Itch mediates the degradation of the p63 protein. Itch contains four WW domains which are pivotal for the substrate recognition process. Indeed, this domain is implicated in several signalling complexes crucially involved in human diseases including Muscular Dystrophy, Alzheimer's Disease and Huntington Disease. WW domains are highly compact protein-protein binding modules that interact with short proline-rich sequences. The four WW domains present in Itch belong to the Group I type, which binds polypeptides with a PY motif characterized by a PP xY consensus sequence, where x can be any residue. Accordingly, the Itch-p63 interaction results from a direct binding of Itch-WW2 domain with the PY motif of p63. Here, we report a structural analysis of the Itch-p63 interaction by fluorescence, CD and NMR spectroscopy. Indeed, we studied the in vitro interaction between Itch-WW2 domain and p63(534-551), an 18-mer peptide encompassing a fragment of the p63 protein including the PY motif. In addition, we evaluated the conformation and the interaction with Itch-WW2 of a site specific mutant of p63, I549T, that has been reported in both Hay-Wells syndrome and Rapp-Hodgkin syndrome. Based on our results, we propose an extended PP xY motif for the Itch recognition motif (P-P-P-Y-x(4)-[ST]-[ILV]), which includes these C-terminal residues to the PP xY motif.
E3 泛素连接酶 Itch 介导 p63 蛋白的降解。Itch 包含四个 WW 结构域，对于底物识别过程至关重要。事实上，该结构域与几种信号复合物有关，这些复合物在包括肌肉萎缩症、阿尔茨海默病和亨廷顿病在内的人类疾病中起着关键作用。WW 结构域是高度紧凑的蛋白质-蛋白质结合模块，与富含脯氨酸的短序列相互作用。Itch 中存在的四个 WW 结构域属于 I 型组，与具有 PY 基序的多肽结合，其特征是 PP xY 一致序列，其中 x 可以是任何残基。因此，Itch-p63 相互作用是由 Itch-WW2 结构域与 p63 的 PY 基序的直接结合引起的。在这里，我们通过荧光、CD 和 NMR 光谱学报告了 Itch-p63 相互作用的结构分析。事实上，我们研究了 Itch-WW2 结构域与 p63(534-551)之间的体外相互作用，p63(534-551)是一个 18 肽，包含 p63 蛋白的一个片段，包括 PY 基序。此外，我们评估了 p63 的一个特定突变体 I549T 的构象和与 Itch-WW2 的相互作用，该突变体已在 Hay-Wells 综合征和 Rapp-Hodgkin 综合征中报道。基于我们的结果，我们提出了一个扩展的 PP xY 基序用于 Itch 识别基序（P-P-P-Y-x(4)-[ST]-[ILV]），该基序包括这些 C 末端残基到 PP xY 基序。
