Poxviruses encode multiple proteins that enable them to evade host responses. Among these are serine protease inhibitors (serpins). One of the earliest serpins described, cowpox virus crmA, acts to inhibit inflammation and apoptosis. crmA homologous serpins, known as SPI-2, are conserved in rabbitpox, vaccinia and variola viruses. Here, we describe the characterization of ectromelia virus (EV) SPI-2. EV SPI-2 encodes a protein of approximately 38 kDa showing >94% identity with other poxviral homologues. Conservative changes in amino acid sequence were found within the reactive site loop and the serpin backbone. Like crmA, transient expression of SPI-2 protected cells from tumour necrosis factor-mediated apoptosis and inhibited the activity of caspases-1 and -8 but not caspases-3, -6 or granzyme B. Overall, this study demonstrates that EV SPI-2 is functionally similar to crmA, based on in vitro assays.