Interaction between human positive coactivator 4 (PC4), an abundant nuclear protein, and the tumor suppressor protein p53 plays a crucial role in initiating apoptosis. In certain neurodegenerative diseases PC4 assisted-p53-dependent apoptosis may play a central role. Thus, disruption of p53-PC4 interaction may be a good drug target for certain disease pathologies. A p53-derived short peptide (AcPep) that binds the C-terminal domain of PC4 (C-PC4) is known to disrupt PC4-p53 interaction. To fully characterize its binding mode and binding site on PC4, we co-crystallized C-PC4 with the peptide and determined its structure. The crystal, despite exhibiting mass spectrometric signature of the peptide, lacked peptide electron density and showed a novel crystal lattice, when compared to C-PC4 crystals without the peptide. Using peptide-docked models of crystal lattices, corresponding to our structure and the peptide-devoid structure we show the origin of the novel crystal lattice to be dynamically bound peptide at the previously identified putative binding site. The weak binding is proposed to be due to the lack of the N-terminal domain of PC4 (N-PC4), which we experimentally show to be disordered with no effect on PC4 stability. Taking cue from the structure, virtual screening of ∼18.6 million small molecules from the ZINC15 database was performed, followed by toxicity and binding free energy filtering. The novel crystal lattice of C-PC4 in presence of the peptide, the role of the disordered N-PC4 and the high throughput identification of potent small molecules will allow a better understanding and control of p53-PC4 interaction.

人类阳性共激活因子 4 (PC4) 是一种丰富的核蛋白，与肿瘤抑制蛋白 p53 之间的相互作用在启动细胞凋亡中起着至关重要的作用。在某些神经退行性疾病中，PC4 辅助的 p53 依赖性细胞凋亡可能发挥核心作用。因此，p53-PC4 相互作用的破坏可能是某些疾病病理的良好药物靶点。p53 衍生的短肽 ( AcPep) 结合 PC4 (C-PC4) 的 C 端结构域已知会破坏 PC4-p53 相互作用。为了充分表征其在 PC4 上的结合模式和结合位点，我们将 C-PC4 与肽共结晶并确定其结构。与不含肽的 C-PC4 晶体相比，该晶体尽管表现出肽的质谱特征，但缺乏肽电子密度并显示出新的晶格。使用与我们的结构和无肽结构相对应的晶格的肽对接模型，我们显示新晶格的起源是在先前确定的推定结合位点动态结合肽。弱结合被认为是由于缺乏 PC4 (N-PC4) 的 N 端结构域，我们实验表明它是无序的，对 PC4 稳定性没有影响。从结构中获取线索，对 ZINC15 数据库中的 1860 万个小分子进行虚拟筛选，然后进行毒性和结合自由能过滤。C-PC4 在肽存在下的新晶格、无序 N-PC4 的作用和有效小分子的高通量鉴定将有助于更好地理解和控制 p53-PC4 相互作用。