The recognition and cleavage of gasdermin D (GSDMD) by inflammatory caspases-1, 4, 5, and 11 are essential steps in initiating pyroptosis after inflammasome activation. Previous work has identified cleavage site signatures in substrates such as GSDMD, but it is unclear whether these are the sole determinants for caspase engagement. Here we report the crystal structure of a complex between human caspase-1 and the full-length murine GSDMD. In addition to engagement of the GSDMD N- and C-domain linker by the caspase-1 active site, an anti-parallel β sheet at the caspase-1 L2 and L2′ loops bound a hydrophobic pocket within the GSDMD C-terminal domain distal to its N-terminal domain. This “exosite” interface endows an additional function for the GSDMD C-terminal domain as a caspase-recruitment module besides its role in autoinhibition. Our study thus reveals dual-interface engagement of GSDMD by caspase-1, which may be applicable to other physiological substrates of caspases.

炎症性 caspases-1、4、5 和 11 对 gasdermin D (GSDMD) 的识别和切割是炎性小体激活后启动细胞焦亡的重要步骤。之前的工作已经确定了 GSDMD 等底物中的切割位点特征，但尚不清楚这些是否是半胱天冬酶参与的唯一决定因素。在这里，我们报告了人类 caspase-1 和全长鼠类 GSDMD 之间的复合物的晶体结构。除了 caspase-1 活性位点与 GSDMD N 和 C 域接头的结合外，caspase-1 L2 和 L2' 环上的反平行 β 片层结合了 GSDMD C 末端域远端的疏水口袋到它的 N 端域。这种“外部位点”界面除了在自身抑制中的作用外，还赋予 GSDMD C 末端域作为 caspase 募集模块的附加功能。