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The Vaccinia Virus Superoxide Dismutase-Like Protein (A45R) Is a Virion Component That Is Nonessential for Virus Replication

By Fernando Almazán, David C. Tscharke and Geoffrey L. Smith

Abstract

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 (Mr, of 13,600) with 39% amino acid identity to copper-zinc superoxide dismutase (Cu-Zn SOD). Sequencing of the A45R gene from other orthopoxviruses, here and by others, showed that the protein is highly conserved 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 important for the catalytic activity. The A45R protein was expressed in Escherichia coli, purified, and tested for SOD activity, but neither enzymatic nor inhibitory SOD activity was detected. Additionally, no virus-encoded SOD activity was detected in infected cells or purified virions. A monoclonal antibody 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 infection. Confocal microscopy of VV-infected cells indicated that the A45R protein accumulated predominantly in cytoplasmic viral factories. Electron microscopy and biochemical analyses showed that the A45R protein is incorporated into the virion core. A deletion mutant lacking the majority of the A45R gene and a revertant virus in which the deleted gene was restored were constructed 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 virulence 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 protein described that affects neither virus replication nor virulence

Topics: Replication
Publisher: American Society for Microbiology
Year: 2001
DOI identifier: 10.1128/JVI.75.15.7018-7029.2001
OAI identifier: oai:pubmedcentral.nih.gov:114430
Provided by: PubMed Central
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