Linear ubiquitylation, in which ubiquitin units are covalently linked through N- and C-terminal amino acids, is a unique cellular signaling mechanism. This process is controlled by a single E3 ubiquitin ligase, the linear ubiquitin chain assembly complex (LUBAC), which is composed of three proteins – HOIL-1L, HOIP and SHARPIN. LUBAC is involved in the activation of the canonical NF-κB pathway and has been linked to NF-κB dependent malignancies. In this work, we present HOIP-based stapled alpha-helical peptides designed to inhibit LUBAC through the disruption of the HOIL-1L-HOIP interaction and loss of the functional complex. We find our HOIP peptides to be active LUBAC ubiquitylation inhibitors in vitro, though through interaction with HOIP rather than HOIL. Active peptides were shown to have inhibitory effects on cell viability, reduced NF-κB activity and decreased production of NF-κB related gene products. This work further demonstrates the potential of LUBAC as a therapeutic target and of the use of stapled peptides as inhibitors of protein-protein interactions.

线性泛素化是一种独特的细胞信号传导机制，其中泛素单元通过N和C末端氨基酸共价连接。此过程由单个E3泛素连接酶（线性泛素链组装复合物（LUBAC））控制，该复合物由三种蛋白质-HOIL-1L，HOIP和SHARPIN组成。LUBAC参与规范性NF-κB途径的激活，并与NF-κB依赖性恶性肿瘤有关。在这项工作中，我们介绍了基于HOIP的钉合α-螺旋肽，旨在通过破坏HOIL-1L-HOIP相互作用和功能复合物的丧失来抑制LUBAC。我们发现我们的HOIP肽在体外是活性LUBAC泛素化抑制剂，尽管是通过与HOIP而非HOIL进行交互。活性肽显示出对细胞存活率具有抑制作用，降低了NF-κB活性并降低了NF-κB相关基因产物的产生。这项工作进一步证明了LUBAC作为治疗靶标的潜力以及将钉合肽用作蛋白质-蛋白质相互作用的抑制剂的潜力。