Orthopoxviruses bear in their genomes several genes coding for homologous secreted proteins able to bind tumor necrosis factor. Different species of the genus possess different sets of these tumor necrosis factor-binding proteins. Viriola virus encodes the only one of them named CrmB. Despite sharing high sequence identity, CrmB proteins belonging to distinct orthopoxviral species were shown to significantly differ by their physico-chemical and biological properties. We modeled spatial structures of tumor necrosis factor receptor domains of variola and cowpox virus CrmB proteins bound to either murine, or human or mutated human tumor necrosis factor. In the sequence of last the arginine residue at position 31 is substituted with glutamine that is characteristic for murine tumor necrosis factor. Theoretical analysis of modeled ligand-receptor complexes revealed that the least stable should be the complex of cowpox virus CrmB with human tumor necrosis factor, and that arginine to glutamine substitution at position 31 should significantly stabilize binding of corresponding human tumor necrosis factor mutant to cowpox virus CrmB. Experimental evaluation of recombinant variola and cowpox virus CrmB efficiencies in inhibiting cytotoxic effect of all these tumor necrosis factors have approved our predictions.