Protein-Protein Interaction Laboratory, The Francis Crick Institute, London, NW1 1AT, UK.
Epigenetics, The Babraham Institute, Cambridge, CB22 3AT, UK.
Nat Commun. 2024 Nov 11;15(1):9746. doi: 10.1038/s41467-024-53554-1.
Peptidylarginine deiminase IV (PADI4, PAD4) deregulation promotes the development of autoimmunity, cancer, atherosclerosis and age-related tissue fibrosis. PADI4 additionally mediates immune responses and cellular reprogramming, although the full extent of its physiological roles is unexplored. Despite detailed molecular knowledge of PADI4 activation in vitro, we lack understanding of its regulation within cells, largely due to a lack of appropriate systems and tools. Here, we develop and apply a set of potent and selective PADI4 modulators. Using the mRNA-display-based RaPID system, we screen >10 cyclic peptides for high-affinity, conformation-selective binders. We report PADI4_3, a cell-active inhibitor specific for the active conformation of PADI4; PADI4_7, an inert binder, which we functionalise for the isolation and study of cellular PADI4; and PADI4_11, a cell-active PADI4 activator. Structural studies with PADI4_11 reveal an allosteric binding mode that may reflect the mechanism that promotes cellular PADI4 activation. This work contributes to our understanding of PADI4 regulation and provides a toolkit for the study and modulation of PADI4 across (patho)physiological contexts.
肽基精氨酸脱亚氨酶 IV（PADI4，PAD4）失调会促进自身免疫、癌症、动脉粥样硬化和与年龄相关的组织纤维化的发展。PADI4 还介导免疫反应和细胞重编程，尽管其生理作用的全部范围尚未得到探索。尽管我们对 PADI4 在体外的激活有详细的分子知识，但我们对其在细胞内的调节知之甚少，这主要是因为缺乏合适的系统和工具。在这里，我们开发并应用了一组有效的、选择性的 PADI4 调节剂。我们使用基于 mRNA 展示的 RaPID 系统筛选了 >10 个环肽，以获得高亲和力、构象选择性的结合物。我们报告了 PADI4_3，这是一种针对 PADI4 活性构象的细胞活性抑制剂；PADI4_7，一种惰性结合物，我们对其进行功能化，用于分离和研究细胞内 PADI4；以及 PADI4_11，一种细胞活性的 PADI4 激活剂。与 PADI4_11 的结构研究揭示了一种别构结合模式，可能反映了促进细胞内 PADI4 激活的机制。这项工作有助于我们理解 PADI4 的调节，并为在（病理）生理环境中研究和调节 PADI4 提供了一个工具包。
