title: Properties of the recombinant TNF-binding proteins from variola, monkeypox, and cowpox viruses are different.
abstract: Tumor necrosis factor (TNF), a potent proinflammatory and antiviral cytokine, is a critical extracellular immune regulator targeted by poxviruses through the activity of virus-encoded family of TNF-binding proteins (CrmB, CrmC, CrmD, and CrmE). The only TNF-binding protein from variola virus (VARV), the causative agent of smallpox, infecting exclusively humans, is CrmB. Here we have aligned the amino acid sequences of CrmB proteins from 10 VARV, 14 cowpox virus (CPXV), and 22 monkeypox virus (MPXV) strains. Sequence analyses demonstrated a high homology of these proteins. The regions homologous to cd00185 domain of the TNF receptor family, determining the specificity of ligand-receptor binding, were found in the sequences of CrmB proteins. In addition, a comparative analysis of the C-terminal SECRET domain sequences of CrmB proteins was performed. The differences in the amino acid sequences of these domains characteristic of each particular orthopoxvirus species were detected. It was assumed that the species-specific distinctions between the CrmB proteins might underlie the differences in these physicochemical and biological properties. The individual recombinant proteins VARV-CrmB, MPXV-CrmB, and CPXV-CrmB were synthesized in a baculovirus expression system in insect cells and isolated. Purified VARV-CrmB was detectable as a dimer with a molecular weight of 90 kDa, while MPXV- and CPXV-CrmBs, as monomers when fractioned by non-reducing SDS-PAGE. The CrmB proteins of VARV, MPXV, and CPXV differed in the efficiencies of inhibition of the cytotoxic effects of human, mouse, or rabbit TNFs in L929 mouse fibroblast cell line. Testing of CrmBs in the experimental model of LPS-induced shock using SPF BALB/c mice detected a pronounced protective effect of VARV-CrmB. Thus, our data demonstrated the difference in anti-TNF activities of VARV-, MPXV-, and CPXV-CrmBs and efficiency of VARV-CrmB rather than CPXV- or MPXV-CrmBs against LPS-induced mortality in mice.
authors.affiliation: State Research Center of Virology and Biotechnology Vector, Koltsovo, Novosibirsk oblast, 630559 Russia.
authors.forename: Irina P
authors.initials: IP
authors.lastname: Gileva
authors.forename: Tatiana S
authors.initials: TS
authors.lastname: Nepomnyashchikh
authors.forename: Denis V
authors.initials: DV
authors.lastname: Antonets
authors.forename: Leonid R
authors.initials: LR
authors.lastname: Lebedev
authors.forename: Galina V
authors.initials: GV
authors.lastname: Kochneva
authors.forename: Antonina V
authors.initials: AV
authors.lastname: Grazhdantseva
authors.forename: Sergei N
authors.initials: SN
authors.lastname: Shchelkunov
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D015608:Cowpox virus / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D008993:Monkeypox virus / Q000378:metabolism*; D018124:Receptors, Tumor Necrosis Factor / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D018411:Spodoptera; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D012901:Variola virus / Q000378:metabolism*; D014764:Viral Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D018124:Receptors, Tumor Necrosis Factor; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D014764:Viral Proteins