The vaccinia virus superoxide dismutase-like protein (A45R) is a virion component that is nonessential for virus replication

A characterization of the A45R gene from vaccinia virus (VV) strain Western Reserve is presented. The open reading frame is predicted to encode a 125- amino-acid protein (M-r, of 13,600) with 39% amino acid identity to copper- zinc superoxide dismutase (Cu-Zn Son), Sequencing of the A45R gene from oth er orthopoxviruses, here and by others, showed that the protein is highly c onserved in all viruses sequenced, including 16 strains of VV, 2 strains of cowpox virus, camelpox virus, and 4 strains of variola virus. In all cases the protein lacks key residues involved in metal ion binding that are impo rtant for the catalytic activity, The A45R protein was expressed in Escheri chia coli, purified, and tested for Son activity, but neither enzymatic nor inhibitory SOD activity was detected, Additionally, no virus-encoded SOD a ctivity was detected in infected cells or purified virions. A monoclonal an tibody raised against the A45R protein expressed in E, coli identified the A45R gene product as a 13.5-kDa protein that is expressed late during VV in fection, Confocal microscopy of VV-infected cells indicated that the A45R p rotein accumulated predominantly in cytoplasmic viral factories, Electron m icroscopy and biochemical analyses showed that the A45R protein is incorpor ated into the virion core. A deletion mutant lacking the majority of the A4 5R gene and a revertant virus in which the deleted gene was restored were c onstructed and characterized. The growth properties of the deletion mutant virus were indistinguishable from those of wild-type and revertant viruses in all cell lines tested, including macrophages, Additionally, the virulenc e and pathogenicity of the three viruses were also comparable in murine and rabbit models of infection, A45R is unusual in being the first VV core pro tein described that affects neither virus replication nor virulence.