IDIBE, Universidad Miguel Hernández, Elche, Spain.
Centro de Biotecnología, Universidad Nacional de Loja, Avda, Loja, Ecuador.
Protein Sci. 2023 Aug;32(8):e4723. doi: 10.1002/pro.4723.
PADI4 is one of the human isoforms of a family of enzymes implicated in the conversion of arginine to citrulline. MDM2 is an E3 ubiquitin ligase which is crucial for down-regulation of degradation of the tumor suppressor gene p53. Given the relationship between both PADI4 and MDM2 with p53-signaling pathways, we hypothesized they may interact directly, and this interaction could be relevant in the context of cancer. Here, we showed their association in the nucleus and cytosol in several cancer cell lines. Furthermore, binding was hampered in the presence of GSK484, an enzymatic PADI4 inhibitor, suggesting that MDM2 could bind to the active site of PADI4, as confirmed by in silico experiments. In vitro and in silico studies showed that the isolated N-terminal region of MDM2, N-MDM2, interacted with PADI4, and residues Thr26, Val28, Phe91 and Lys98 were more affected by the presence of the enzyme. Moreover, the dissociation constant between N-MDM2 and PADI4 was comparable to the IC of GSK484 from in cellulo experiments. The interaction between MDM2 and PADI4 might imply MDM2 citrullination, with potential therapeutic relevance for improving cancer treatment, due to the generation of new antigens.
PADI4 是参与精氨酸向瓜氨酸转化的酶家族的人类同工酶之一。MDM2 是一种 E3 泛素连接酶，对于下调肿瘤抑制基因 p53 的降解至关重要。鉴于 PADI4 和 MDM2 与 p53 信号通路之间的关系，我们假设它们可能直接相互作用，这种相互作用在癌症背景下可能是相关的。在这里，我们在几种癌细胞系中显示了它们在核和细胞质中的关联。此外，在存在 PADI4 酶抑制剂 GSK484 的情况下，结合受到阻碍，这表明 MDM2 可以结合到 PADI4 的活性位点，这通过计算机模拟实验得到了证实。体外和计算机模拟研究表明，MDM2 的 N 端区域（N-MDM2）与 PADI4 相互作用，并且残基 Thr26、Val28、Phe91 和 Lys98 受酶的存在影响更大。此外，N-MDM2 和 PADI4 之间的解离常数与细胞内实验中 GSK484 的 IC 相当。MDM2 和 PADI4 之间的相互作用可能意味着 MDM2 的瓜氨酸化，由于产生新的抗原，这可能对改善癌症治疗具有潜在的治疗意义。
