Department of Pediatrics, Suzhou Kowloon Hospital, Shanghai Jiao Tong University School of Medicine, Suzhou, China.
J Mol Recognit. 2023 Aug;36(8):e3045. doi: 10.1002/jmr.3045. Epub 2023 Jul 6.
Interleukin-17A (IL-17A) is a pro-inflammatory cytokine implicated in diverse autoimmune and inflammatory disorders such as psoriasis and Kawasaki disease. Mature IL-17A is a homodimer that binds to the extracellular type-III fibronectin D1:D2-dual domain of its cognate IL-17 receptor A (IL-17RA). In this study, we systematically examined the structural basis, thermodynamics property, and dynamics behavior of IL-17RA/IL-17A interaction and computationally identified two continuous hotspot regions separately from different monomers of IL-17A homodimer that contribute significantly to the interaction, namely I-shaped and U-shaped segments, thus rendered as a peptide-mediated protein-protein interaction (PmPPI). Self-inhibitory peptides (SIPs) are derived from the two segments to disrupt IL-17RA/IL-17A interaction by competitively rebinding to the IL-17A-binding pocket on IL-17RA surface, which, however, only have a weak affinity and low specificity for IL-17RA due to lack of the context support of intact IL-17A protein, thus exhibiting a large flexibility and intrinsic disorder when splitting from the protein context and incurring a considerable entropy penalty when rebinding to IL-17RA. The U-shaped segment is further extended, mutated and stapled by a disulfide bridge across its two strands to obtain a number of double-stranded cyclic SIPs, which are partially ordered and conformationally similar to their native status at IL-17RA/IL-17A complex interface. Experimental fluorescence polarization assays substantiate that the stapling can moderately or considerably improve the binding affinity of U-shaped segment-derived peptides by 2-5-fold. In addition, computational structural modeling also reveals that the stapled peptides can bind in a similar mode with the native crystal conformation of U-shaped segment in IL-17RA pocket, where the disulfide bridge is out of the pocket for avoiding intervene of the peptide binding.
白细胞介素-17A (IL-17A) 是一种促炎细胞因子，与多种自身免疫和炎症性疾病有关，如银屑病和川崎病。成熟的 IL-17A 是一种同二聚体，与它的同源性白细胞介素-17 受体 A (IL-17RA) 的细胞外 III 型纤维连接蛋白 D1:D2 双域结合。在这项研究中，我们系统地研究了 IL-17RA/IL-17A 相互作用的结构基础、热力学性质和动力学行为，并从 IL-17A 同二聚体的两个不同单体中计算出分别对相互作用有重要贡献的两个连续热点区域，即 I 形和 U 形片段，从而构成肽介导的蛋白质-蛋白质相互作用（PmPPI）。自抑制肽（SIP）来源于这两个片段，通过竞争性地与 IL-17RA 表面的 IL-17A 结合口袋重新结合来破坏 IL-17RA/IL-17A 相互作用，但由于缺乏完整的 IL-17A 蛋白的上下文支持，SIP 对 IL-17RA 只有较弱的亲和力和较低的特异性，因此当从蛋白质环境中分裂时表现出较大的灵活性和固有无序性，并在重新与 IL-17RA 结合时产生相当大的熵罚。U 形片段进一步延伸、突变并通过二硫键在其两条链上交错，得到一些双链环状 SIP，这些 SIP 部分有序且构象与它们在 IL-17RA/IL-17A 复合物界面上的天然状态相似。荧光偏振实验证实，交联可以适度或显著提高 U 形片段衍生肽的结合亲和力 2-5 倍。此外，计算结构建模还揭示了，交联肽可以以与 U 形片段在 IL-17RA 口袋中的天然晶体构象相似的方式结合，其中二硫键不在口袋内，以避免肽结合的干扰。
