Abstract
The vaccinia virus D5 gene encodes a 90-kDa protein that is transiently expressed at early times after infection. Temperature-sensitive mutants with lesions in the D5 gene exhibit a fast-stop DNA- phenotype and are also impaired in homologous recombination. Here we report the overexpression of the D5 protein within the context of a vaccinia virus infection and its purification to apparent homogeneity. The purified protein has an intrinsic nucleoside triphosphatase activity which is independent of, and not stimulated by, any common nucleic acid cofactors. All eight common ribo- and deoxyribonucleoside triphosphates are hydrolyzed to the diphosphate form in the presence of a divalent cation. Implications for the role of D5 in viral DNA replication are addressed.
Full Text
The Full Text of this article is available as a PDF (505.3 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Alexander W. A., Moss B., Fuerst T. R. Regulated expression of foreign genes in vaccinia virus under the control of bacteriophage T7 RNA polymerase and the Escherichia coli lac repressor. J Virol. 1992 May;66(5):2934–2942. doi: 10.1128/jvi.66.5.2934-2942.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Arai K., Kornberg A. Mechanism of dnaB protein action. II. ATP hydrolysis by dnaB protein dependent on single- or double-stranded DNA. J Biol Chem. 1981 May 25;256(10):5253–5259. [PubMed] [Google Scholar]
- Broyles S. S., Yuen L., Shuman S., Moss B. Purification of a factor required for transcription of vaccinia virus early genes. J Biol Chem. 1988 Aug 5;263(22):10754–10760. [PubMed] [Google Scholar]
- Chen M., Pan Z. Q., Hurwitz J. Sequence and expression in Escherichia coli of the 40-kDa subunit of activator 1 (replication factor C) of HeLa cells. Proc Natl Acad Sci U S A. 1992 Apr 1;89(7):2516–2520. doi: 10.1073/pnas.89.7.2516. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Condit R. C., Motyczka A. Isolation and preliminary characterization of temperature-sensitive mutants of vaccinia virus. Virology. 1981 Aug;113(1):224–241. doi: 10.1016/0042-6822(81)90150-1. [DOI] [PubMed] [Google Scholar]
- Condit R. C., Motyczka A., Spizz G. Isolation, characterization, and physical mapping of temperature-sensitive mutants of vaccinia virus. Virology. 1983 Jul 30;128(2):429–443. doi: 10.1016/0042-6822(83)90268-4. [DOI] [PubMed] [Google Scholar]
- De-Medina T., Shaul Y. Functional and structural similarity between the X protein of hepatitis B virus and nucleoside diphosphate kinases. FEBS Lett. 1994 Sep 12;351(3):423–426. doi: 10.1016/0014-5793(94)00900-7. [DOI] [PubMed] [Google Scholar]
- Elroy-Stein O., Fuerst T. R., Moss B. Cap-independent translation of mRNA conferred by encephalomyocarditis virus 5' sequence improves the performance of the vaccinia virus/bacteriophage T7 hybrid expression system. Proc Natl Acad Sci U S A. 1989 Aug;86(16):6126–6130. doi: 10.1073/pnas.86.16.6126. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ensinger M. J. Isolation and genetic characterization of temperature-sensitive mutants of vaccinia virus WR. J Virol. 1982 Sep;43(3):778–790. doi: 10.1128/jvi.43.3.778-790.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ensinger M. J., Rovinsky M. Marker rescue of temperature-sensitive mutations of vaccinia virus WR: correlation of genetic and physical maps. J Virol. 1983 Nov;48(2):419–428. doi: 10.1128/jvi.48.2.419-428.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Evans E., Traktman P. Characterization of vaccinia virus DNA replication mutants with lesions in the D5 gene. Chromosoma. 1992;102(1 Suppl):S72–S82. doi: 10.1007/BF02451789. [DOI] [PubMed] [Google Scholar]
- Evans E., Traktman P. Molecular genetic analysis of a vaccinia virus gene with an essential role in DNA replication. J Virol. 1987 Oct;61(10):3152–3162. doi: 10.1128/jvi.61.10.3152-3162.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gershon P. D., Ahn B. Y., Garfield M., Moss B. Poly(A) polymerase and a dissociable polyadenylation stimulatory factor encoded by vaccinia virus. Cell. 1991 Sep 20;66(6):1269–1278. doi: 10.1016/0092-8674(91)90048-4. [DOI] [PubMed] [Google Scholar]
- Gorbalenya A. E., Koonin E. V. Viral proteins containing the purine NTP-binding sequence pattern. Nucleic Acids Res. 1989 Nov 11;17(21):8413–8440. doi: 10.1093/nar/17.21.8413. [DOI] [PMC free article] [PubMed] [Google Scholar]
- JOKLIK W. K., BECKER Y. THE REPLICATION AND COATING OF VACCINIA DNA. J Mol Biol. 1964 Dec;10:452–474. doi: 10.1016/s0022-2836(64)80066-8. [DOI] [PubMed] [Google Scholar]
- Johnson G. P., Goebel S. J., Paoletti E. An update on the vaccinia virus genome. Virology. 1993 Oct;196(2):381–401. doi: 10.1006/viro.1993.1494. [DOI] [PubMed] [Google Scholar]
- Jong A. Y., Campbell J. L. Characterization of Saccharomyces cerevisiae thymidylate kinase, the CDC8 gene product. General properties, kinetic analysis, and subcellular localization. J Biol Chem. 1984 Dec 10;259(23):14394–14398. [PubMed] [Google Scholar]
- Jong A. Y., Ma J. J. Saccharomyces cerevisiae nucleoside-diphosphate kinase: purification, characterization, and substrate specificity. Arch Biochem Biophys. 1991 Dec;291(2):241–246. doi: 10.1016/0003-9861(91)90129-7. [DOI] [PubMed] [Google Scholar]
- Kerr S. M., Smith G. L. Vaccinia virus encodes a polypeptide with DNA ligase activity. Nucleic Acids Res. 1989 Nov 25;17(22):9039–9050. doi: 10.1093/nar/17.22.9039. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kim H. J., Nishikawa S., Tokutomi Y., Takenaka H., Hamada M., Kuby S. A., Uesugi S. In vitro mutagenesis studies at the arginine residues of adenylate kinase. A revised binding site for AMP in the X-ray-deduced model. Biochemistry. 1990 Feb 6;29(5):1107–1111. doi: 10.1021/bi00457a002. [DOI] [PubMed] [Google Scholar]
- Koonin E. V. A common set of conserved motifs in a vast variety of putative nucleic acid-dependent ATPases including MCM proteins involved in the initiation of eukaryotic DNA replication. Nucleic Acids Res. 1993 Jun 11;21(11):2541–2547. doi: 10.1093/nar/21.11.2541. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Koonin E. V., Senkevich T. G. Vaccinia virus encodes four putative DNA and/or RNA helicases distantly related to each other. J Gen Virol. 1992 Apr;73(Pt 4):989–993. doi: 10.1099/0022-1317-73-4-989. [DOI] [PubMed] [Google Scholar]
- Lascu I., Deville-Bonne D., Glaser P., Véron M. Equilibrium dissociation and unfolding of nucleoside diphosphate kinase from Dictyostelium discoideum. Role of proline 100 in the stability of the hexameric enzyme. J Biol Chem. 1993 Sep 25;268(27):20268–20275. [PubMed] [Google Scholar]
- Mackett M., Smith G. L., Moss B. General method for production and selection of infectious vaccinia virus recombinants expressing foreign genes. J Virol. 1984 Mar;49(3):857–864. doi: 10.1128/jvi.49.3.857-864.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Mathews C. K. Enzyme organization in DNA precursor biosynthesis. Prog Nucleic Acid Res Mol Biol. 1993;44:167–203. doi: 10.1016/s0079-6603(08)60220-2. [DOI] [PubMed] [Google Scholar]
- McDonald W. F., Traktman P. Overexpression and purification of the vaccinia virus DNA polymerase. Protein Expr Purif. 1994 Aug;5(4):409–421. doi: 10.1006/prep.1994.1059. [DOI] [PubMed] [Google Scholar]
- McFadden G., Dales S. Biogenesis of poxviruses: preliminary characterization of conditional lethal mutants of vaccinia virus defective in DNA synthesis. Virology. 1980 May;103(1):68–79. doi: 10.1016/0042-6822(80)90126-9. [DOI] [PubMed] [Google Scholar]
- Mimouni M., Bontemps F., Van den Berghe G. Kinetic studies of rat liver adenosine kinase. Explanation of exchange reaction between adenosine and AMP. J Biol Chem. 1994 Jul 8;269(27):17820–17825. [PubMed] [Google Scholar]
- Morgan J. R., Cohen L. K., Roberts B. E. Identification of the DNA sequences encoding the large subunit of the mRNA-capping enzyme of vaccinia virus. J Virol. 1984 Oct;52(1):206–214. doi: 10.1128/jvi.52.1.206-214.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Moss B., Elroy-Stein O., Mizukami T., Alexander W. A., Fuerst T. R. Product review. New mammalian expression vectors. Nature. 1990 Nov 1;348(6296):91–92. doi: 10.1038/348091a0. [DOI] [PubMed] [Google Scholar]
- Neuhard J., Randerath E., Randerath K. Ion-exchange thin-layer chromatography. 13. Resolution of complex nucleoside triphosphate mixtures. Anal Biochem. 1965 Nov;13(2):211–222. doi: 10.1016/0003-2697(65)90191-0. [DOI] [PubMed] [Google Scholar]
- Niles E. G., Condit R. C., Caro P., Davidson K., Matusick L., Seto J. Nucleotide sequence and genetic map of the 16-kb vaccinia virus HindIII D fragment. Virology. 1986 Aug;153(1):96–112. doi: 10.1016/0042-6822(86)90011-5. [DOI] [PubMed] [Google Scholar]
- Paolette E., Rosemond-Hornbeak H., Moss B. Two nucleid acid-dependent nucleoside triphosphate phosphohydrolases from vaccinia virus. Purification and characterization. J Biol Chem. 1974 May 25;249(10):3273–3280. [PubMed] [Google Scholar]
- Paoletti E., Moss B. Two nucleic acid-dependent nucleoside triphosphate phosphohydrolases from vaccinia virus. Nucleotide substrate and polynucleotide cofactor specificities. J Biol Chem. 1974 May 25;249(10):3281–3286. [PubMed] [Google Scholar]
- Pelham H. R., Jackson R. J. An efficient mRNA-dependent translation system from reticulocyte lysates. Eur J Biochem. 1976 Aug 1;67(1):247–256. doi: 10.1111/j.1432-1033.1976.tb10656.x. [DOI] [PubMed] [Google Scholar]
- Rempel R. E., Anderson M. K., Evans E., Traktman P. Temperature-sensitive vaccinia virus mutants identify a gene with an essential role in viral replication. J Virol. 1990 Feb;64(2):574–583. doi: 10.1128/jvi.64.2.574-583.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Roseman N. A., Hruby D. E. Nucleotide sequence and transcript organization of a region of the vaccinia virus genome which encodes a constitutively expressed gene required for DNA replication. J Virol. 1987 May;61(5):1398–1406. doi: 10.1128/jvi.61.5.1398-1406.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Seo Y. S., Hurwitz J. Isolation of helicase alpha, a DNA helicase from HeLa cells stimulated by a fork structure and signal-stranded DNA-binding proteins. J Biol Chem. 1993 May 15;268(14):10282–10295. [PubMed] [Google Scholar]
- Shimizu K., Sugino A. Purification and characterization of DNA helicase III from the yeast Saccharomyces cerevisiae. J Biol Chem. 1993 May 5;268(13):9578–9584. [PubMed] [Google Scholar]
- Shuman S. Vaccinia virus RNA helicase. Directionality and substrate specificity. J Biol Chem. 1993 Jun 5;268(16):11798–11802. [PubMed] [Google Scholar]
- Shuman S. Vaccinia virus RNA helicase: an essential enzyme related to the DE-H family of RNA-dependent NTPases. Proc Natl Acad Sci U S A. 1992 Nov 15;89(22):10935–10939. doi: 10.1073/pnas.89.22.10935. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Siegel L. M., Monty K. J. Determination of molecular weights and frictional ratios of proteins in impure systems by use of gel filtration and density gradient centrifugation. Application to crude preparations of sulfite and hydroxylamine reductases. Biochim Biophys Acta. 1966 Feb 7;112(2):346–362. doi: 10.1016/0926-6585(66)90333-5. [DOI] [PubMed] [Google Scholar]
- Smith G. L., Chan Y. S., Howard S. T. Nucleotide sequence of 42 kbp of vaccinia virus strain WR from near the right inverted terminal repeat. J Gen Virol. 1991 Jun;72(Pt 6):1349–1376. doi: 10.1099/0022-1317-72-6-1349. [DOI] [PubMed] [Google Scholar]
- Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
- Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Traktman P., Kelvin M., Pacheco S. Molecular genetic analysis of vaccinia virus DNA polymerase mutants. J Virol. 1989 Feb;63(2):841–846. doi: 10.1128/jvi.63.2.841-846.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Traktman P., Sridhar P., Condit R. C., Roberts B. E. Transcriptional mapping of the DNA polymerase gene of vaccinia virus. J Virol. 1984 Jan;49(1):125–131. doi: 10.1128/jvi.49.1.125-131.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Traktman P. The enzymology of poxvirus DNA replication. Curr Top Microbiol Immunol. 1990;163:93–123. doi: 10.1007/978-3-642-75605-4_4. [DOI] [PubMed] [Google Scholar]
- Tsurimoto T., Stillman B. Functions of replication factor C and proliferating-cell nuclear antigen: functional similarity of DNA polymerase accessory proteins from human cells and bacteriophage T4. Proc Natl Acad Sci U S A. 1990 Feb;87(3):1023–1027. doi: 10.1073/pnas.87.3.1023. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Walker J. E., Saraste M., Runswick M. J., Gay N. J. Distantly related sequences in the alpha- and beta-subunits of ATP synthase, myosin, kinases and other ATP-requiring enzymes and a common nucleotide binding fold. EMBO J. 1982;1(8):945–951. doi: 10.1002/j.1460-2075.1982.tb01276.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Weiner B. M., Bradley M. K. Specific mutation of a regulatory site within the ATP-binding region of simian virus 40 large T antigen. J Virol. 1991 Sep;65(9):4973–4984. doi: 10.1128/jvi.65.9.4973-4984.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Yoder B. L., Burgers P. M. Saccharomyces cerevisiae replication factor C. I. Purification and characterization of its ATPase activity. J Biol Chem. 1991 Nov 25;266(33):22689–22697. [PubMed] [Google Scholar]
- Zhu L. A., Weller S. K. The UL5 gene of herpes simplex virus type 1: isolation of a lacZ insertion mutant and association of the UL5 gene product with other members of the helicase-primase complex. J Virol. 1992 Jan;66(1):458–468. doi: 10.1128/jvi.66.1.458-468.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zhu L., Weller S. K. UL5, a protein required for HSV DNA synthesis: genetic analysis, overexpression in Escherichia coli, and generation of polyclonal antibodies. Virology. 1988 Oct;166(2):366–378. doi: 10.1016/0042-6822(88)90507-7. [DOI] [PubMed] [Google Scholar]
- de-Medina T., Haviv I., Noiman S., Shaul Y. The X protein of hepatitis B virus has a ribo/deoxy ATPase activity. Virology. 1994 Jul;202(1):401–407. doi: 10.1006/viro.1994.1356. [DOI] [PubMed] [Google Scholar]