Abstract:
:The D4R gene of vaccinia virus encodes a functional uracil-DNA glycosylase that is essential for viral viability (D. T. Stuart, C. Upton, M. A. Higman, E. G. Niles, and G. McFadden, J. Virol. 67:2503-2513, 1993), and a D4R mutant, ts4149, confers a conditional lethal defect in viral DNA replication (A. K. Millns, M. S. Carpenter, and A. M. DeLange, Virology 198:504-513, 1994). The mutant ts4149 protein was expressed in vitro and assayed for uracil-DNA glycosylase activity. Less than 6% of wild-type activity was observed at permissive temperatures, but the ts4149 protein was completely inactive at the nonpermissive temperature. Mutagenesis of the ts4149 gene back to wild type (Arg-179-->Gly) restored full activity. The ts4149 protein was considerably reduced in lysates of cells infected at the permissive temperature, and its activity was undetectable, even in the presence of the uracil glycosylase inhibitor protein, which inhibits the host uracil-DNA glycosylases but not that of vaccinia virus. Thus the ts4149 protein is thermolabile, correlating uracil removal with vaccinia virus DNA replication. Three active-site amino acids of the vaccinia virus uracil-DNA glycosylase were mutated (Asp-68-->Asn, Asn-120-->Val, and His-181-->Leu), producing proteins that were completely defective in uracil excision but still retained the ability to bind DNA. Each mutated D4R gene was transfected into vaccinia virus ts4149-infected cells in order to assess the recombination events that allowed virus survival at 40 degrees C. Genetic analysis and sequencing studies revealed that the only viruses to survive were those in which recombination eliminated the mutant locus. We conclude that the uracil cleavage activity of the D4R protein is essential for its function in vaccinia virus DNA replication, suggesting that the removal of uracil residues plays an obligatory role.
journal_name
J Viroljournal_title
Journal of virologyauthors
Ellison KS,Peng W,McFadden Gdoi
10.1128/JVI.70.11.7965-7973.1996subject
Has Abstractpub_date
1996-11-01 00:00:00pages
7965-73issue
11eissn
0022-538Xissn
1098-5514journal_volume
70pub_type
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journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.63.12.5194-5200.1989
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doi:10.1128/JVI.13.1.125-139.1974
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doi:10.1128/JVI.65.2.672-676.1991
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doi:10.1128/JVI.66.1.172-182.1992
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pub_type: 杂志文章
doi:10.1128/JVI.71.1.746-749.1997
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journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.32.2.394-403.1979
更新日期:1979-11-01 00:00:00
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doi:10.1128/JVI.62.2.586-593.1988
更新日期:1988-02-01 00:00:00
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pub_type: 杂志文章,评审
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journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.70.10.6557-6562.1996
更新日期:1996-10-01 00:00:00
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journal_title:Journal of virology
pub_type: 杂志文章
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更新日期:2007-11-01 00:00:00
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pub_type: 杂志文章
doi:10.1128/JVI.72.8.6475-6481.1998
更新日期:1998-08-01 00:00:00
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journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.58.2.583-591.1986
更新日期:1986-05-01 00:00:00
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journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.67.11.6469-6475.1993
更新日期:1993-11-01 00:00:00
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journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/jvi.77.9.5178-5191.2003
更新日期:2003-05-01 00:00:00
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journal_title:Journal of virology
pub_type: 杂志文章
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pub_type: 杂志文章
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更新日期:1995-02-01 00:00:00
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journal_title:Journal of virology
pub_type: 杂志文章
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更新日期:2002-08-01 00:00:00
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pub_type: 杂志文章
doi:10.1128/jvi.74.21.10260-10268.2000
更新日期:2000-11-01 00:00:00
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journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.72.4.2638-2646.1998
更新日期:1998-04-01 00:00:00
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pub_type: 杂志文章
doi:10.1128/JVI.2.7.657-662.1968
更新日期:1968-07-01 00:00:00
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journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.45.3.995-1003.1983
更新日期:1983-03-01 00:00:00
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journal_title:Journal of virology
pub_type: 杂志文章
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更新日期:2013-11-01 00:00:00
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journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.67.5.2922-2927.1993
更新日期:1993-05-01 00:00:00