Protocol to purify recombinant inflammatory caspases and assess their catalytic activity in vitro

The inflammatory caspases, such as caspase-1, caspase-4, or caspase-11, are key enzymes in mammalian innate immunity as they control inflammasome-dependent inflammation. Assessing the specific proteolytic activities of these caspases in the context of a cell remains challenging, which is why in vitro studies of their catalytic activity have proven useful. Herein, we describe a detailed protocol for the purification of recombinant inflammatory caspases after heterologous expression in bacteria and how to assess and quantify cleavage of full-length protein substrates. For complete details on the use and execution of this protocol, please refer to Devant et al. (2021).¹.