Synthetic inhibitors of the serine protease DPP9 activate the related NLRP1 and CARD8 inflammasomes and stimulate powerful innate immune responses. Thus, it seems plausible that a biomolecule similarly inhibits DPP9 and triggers inflammasome activation during infection, but one has not yet been discovered. Here, we wanted to identify and characterize DPP9-binding proteins to potentially uncover physiologically relevant mechanisms that control DPP9’s activity. Notably, we found that the redox sensor protein KEAP1 binds to DPP9 in an inactive conformation and stabilizes this non-native fold. At the same time, this inactive form of DPP9 reciprocally inhibits the ability of KEAP1 to bind to and degrade the transcription factor NRF2, thereby inducing an antioxidant response. Although we discovered several experimental conditions, for example new protein expression and chemical denaturation, that force DPP9 out of its folded dimeric state and into a KEAP1-binding state, the key danger-related stimulus that causes this critical DPP9 conformational change is not yet known. Regardless, our data now reveal that an endogenous DPP9 inhibition mechanism does in fact exist, and moreover that DPP9, like the other NLRP1 regulator thioredoxin-1, is directly coupled to the intracellular redox potential. Overall, we expect this work will provide the foundation to discover additional biomolecules that regulate DPP9’s activity, the DPP9-KEAP1 interaction, the intracellular redox environment, and the NLRP1 and CARD8 inflammasomes.

中文翻译：

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丝氨酸蛋白酶 DPP9 和氧化还原传感器 KEAP1 形成相互抑制复合物


丝氨酸蛋白酶 DPP9 的合成抑制剂激活相关的 NLRP1 和 CARD8 炎性小体，并刺激强大的先天免疫反应。因此，生物分子在感染过程中类似地抑制 DPP9 并触发炎性小体激活似乎是合理的，但尚未发现。在这里，我们想鉴定和表征 DPP9 结合蛋白，以潜在地揭示控制 DPP9 活性的生理相关机制。值得注意的是，我们发现氧化还原传感器蛋白 KEAP1 以非活性构象与 DPP9 结合并稳定这种非天然折叠。同时，DPP9 的这种非活性形式相互抑制 KEAP1 结合和降解转录因子 NRF2 的能力，从而诱导抗氧化反应。尽管我们发现了几种实验条件，例如新蛋白表达和化学变性，它们迫使 DPP9 脱离其折叠的二聚体状态并进入 KEAP1 结合状态，但导致这种关键 DPP9 构象变化的关键危险相关刺激尚不清楚。无论如何，我们的数据现在显示内源性 DPP9 抑制机制实际上确实存在，而且 DPP9 与其他 NLRP1 调节因子硫氧还蛋白-1 一样，直接与细胞内氧化还原电位偶联。总体而言，我们预计这项工作将为发现调节 DPP9 活性、DPP9-KEAP1 相互作用、细胞内氧化还原环境以及 NLRP1 和 CARD8 炎性小体的其他生物分子奠定基础。