Cytokine response modifer A (crmA) from the cowpox virus was the first serpin reported to be a �cross-class" inhibitor of both the serine protease, granzyme B, and the cysteine-type proteases, the caspases. Here, we analyze the mechanism by which crmA inhibits these two mechanistically and structurally distinct classes of protease. Engineered single cysteine crmAs labeled with NBD or dansyl fluorophores were found to show similar fluorescence perturbation and FRET changes when crmA formed stable inhibited complexes with either granzyme B or caspase-1. This suggests that crmA inhibits both classes of proteases by a common mechanism in which the formation of a serpin-protease acyl-intermediate complex induces the protease to be translocated to the opposite pole of the serpin and have its catalytic function abolished by conformational deformation of the active-site. However, important mechanistic differences were evident from comparisons of the crm A complexes with granzyme B, a monomer, and caspase-1, a tetramer of two p20p10 catalytic units. Size exclusion chromatography, MALDI-MS, sequencing, native PAGE and Western blotting thus showed that the crmA-caspase-1 complex consisted of a single crmA linked through a thioester bond to a p20 subunit with complete loss of the p10 subunit. Similar findings were made with the complex of crmA with the initiator caspase-8, but the crmA complex with the executioner caspase-6 retained both p20 and p10 subunits. These observations suggest that crmA inhibits serine and cysteine proteases by a similar translocation-distortion mechanism, but that the inhibition of the upstream initiator caspases is made effectively irreversible by the distortion of the protease p20p10 interface as well as the p10p10 interface through which the two caspase catalytic units are associated. The cowpox virus thus exploits the serpin mechanism to provide a novel means of shutting down the proinflammatory and proapoptotic responses of the cell. Grant support: HL78827 and HL79430.