Skip to main content
NCBI home page
Journal of Virology logoLink to Journal of Virology
. 1977 Jul;23(1):53–60. doi: 10.1128/jvi.23.1.53-60.1977

Further Studies on Bacteriophage T4 DNA Synthesis in Sucrose-Plasmolyzed Cells

Mary E Stafford 1, G Prem Veer Reddy 1, Christopher K Mathews 1
PMCID: PMC515799  PMID: 328926

Abstract

This paper describes several technical improvements in the sucrose-plasmolyzed cell system used in earlier experiments on DNA synthesis in situ with Escherichia coli infected by DNA-defective mutants of bacteriophage T4 (W. L. Collinsworth and C. K. Mathews, J. Virol. 13:908-915, 1974). Using this system, which is based primarily on that of M. G. Wovcha et al. (Proc. Natl. Acad. Sci. U.S.A. 70:2196-2200, 1973), we reinvestigated the properties of mutants bearing lesions in genes 1, 41, and 62, and we resolved some disagreements with data reported from that laboratory. We also asked whether the DNA-delay phenotype of T4 mutants is related to possible early leakage of DNA precursors from infected cells. Such cells display defective DNA synthesis in situ, even when ample DNA precursors are made available. Thus, the lesions associated with these mutations seem to manifest themselves at the level of macromolecular metabolism. Similarly, we examined an E. coli mutant defective in its ability to support T4 production, apparently because of a lesion affecting DNA synthesis (L. Simon et al., Nature [London] 252:451-455). In the plasmolyzed cell system, reduced nucleotide incorporation is seen, indicating also that the genetic defect does not involve DNA precursor synthesis. The plasmolyzed cell system incorporates deoxynucleotide 5′-monophosphates into DNA severalfold more rapidly than the corresponding 5′-triphosphates. This is consistent with the idea that DNA precursor-synthesizing enzymes are functionally organized to shuttle substrates to their sites of utilization.

Full text

PDF
53

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Collinsworth W. L., Mathews C. K. Biochemistry of DNA-defective amber mutants of bacteriophage T4. IV. DNA synthesis in plasmolyzed cells. J Virol. 1974 Apr;13(4):908–915. doi: 10.1128/jvi.13.4.908-915.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Dicou L., Cozzarelli N. R. Bacteriophage T4-directed DNA synthesis in toluene-treated cells. J Virol. 1973 Dec;12(6):1293–1302. doi: 10.1128/jvi.12.6.1293-1302.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Duckworth D. H., Bessman M. J. The enzymology of virus-infected bacteria. X. A biochemical-genetic study of the deoxynucleotide kinase induced by wild type and amber mutants of phage T4. J Biol Chem. 1967 Jun 25;242(12):2877–2885. [PubMed] [Google Scholar]
  4. Huang W. M. Membrane-associated proteins of T4-infected Escherichia coli. Virology. 1975 Aug;66(2):508–521. doi: 10.1016/0042-6822(75)90223-8. [DOI] [PubMed] [Google Scholar]
  5. Imae Y., Shinozaki K., Okazaki R. Replication of T4 DNA in vitro. II. Assay system for and some properties of gene products required for T4 DNA replication. J Virol. 1976 Sep;19(3):765–774. doi: 10.1128/jvi.19.3.765-774.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Mathews C. K. Biochemistry of DNA-defective mutants of bacteriophage T4. Thymine nucleotide pool dynamics. Arch Biochem Biophys. 1976 Jan;172(1):178–187. doi: 10.1016/0003-9861(76)90064-3. [DOI] [PubMed] [Google Scholar]
  7. Mathews C. K. Biochemistry of deoxyribonucleic acid-defective amber mutant of bacteriophage T4. I. Ribonucleic acid metabolism. J Biol Chem. 1968 Nov 10;243(21):5610–5615. [PubMed] [Google Scholar]
  8. Mathews C. K. Biochemistry of deoxyribonucleic acid-defective amber mutants of bacteriophage T4. 3. Nucleotide pools. J Biol Chem. 1972 Nov 25;247(22):7430–7438. [PubMed] [Google Scholar]
  9. Matthews C. K. Deoxyribonucleic acid metabolism and virus-induced enzyme synthesis in a thymine-requiring bacterium infected by a thymine-requiring bacteriophage. Biochemistry. 1966 Jun;5(6):2092–2100. doi: 10.1021/bi00870a042. [DOI] [PubMed] [Google Scholar]
  10. McCarthy D., Minner C., Bernstein H., Bernstein C. DNA elongation rates and growing point distributions of wild-type phage T4 and a DNA-delay amber mutant. J Mol Biol. 1976 Oct 5;106(4):963–981. doi: 10.1016/0022-2836(76)90346-6. [DOI] [PubMed] [Google Scholar]
  11. Miller R. C., Jr, Taylor D. M., MacKay K., Smith H. W. Replication of T4 DNA in Escherichia coli treated with toluene. J Virol. 1973 Dec;12(6):1195–1203. doi: 10.1128/jvi.12.6.1195-1203.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Mufti S., Bernstein H. The DNA-delay mutants of bacteriophage T4. J Virol. 1974 Oct;14(4):860–871. doi: 10.1128/jvi.14.4.860-871.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Naot Y., Shalitin C. Role of gene 52 in bacteriophage T4 DNA synthesis. J Virol. 1973 Jun;11(6):862–871. doi: 10.1128/jvi.11.6.862-871.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. North T. W., Stafford M. E., Mathews C. K. Biochemistry of DNA-defective mutants of bacteriophage T4. VI. Biological functions of gene 42. J Virol. 1976 Mar;17(3):973–982. doi: 10.1128/jvi.17.3.973-982.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Simon L. D., Snover D., Doermann A. H. Bacterial mutation affecting T4 phage DNA synthesis and tail production. Nature. 1974 Dec 6;252(5483):451–455. doi: 10.1038/252451a0. [DOI] [PubMed] [Google Scholar]
  16. Tomich P. K., Chiu C. S., Wovcha M. G., Greenberg G. R. Evidence for a complex regulating the in vivo activities of early enzymes induced by bacteriophage T4. J Biol Chem. 1974 Dec 10;249(23):7613–7622. [PubMed] [Google Scholar]
  17. Warner H. R., Hobbs M. D. Incorporation of uracil-14C into nucleic acids in Escherichia coli infected with bacteriophage T4 and T4 amber mutants. Virology. 1967 Nov;33(3):376–384. doi: 10.1016/0042-6822(67)90113-4. [DOI] [PubMed] [Google Scholar]
  18. Weinstein B. I., Bhardwaj N., Li H. C. A rapid and accurate procedure for assaying DNA polymerase, RNA polymerase, or ribosome dependent protein synthesis. Anal Biochem. 1975 Sep;68(1):62–69. doi: 10.1016/0003-2697(75)90679-x. [DOI] [PubMed] [Google Scholar]
  19. Wickner R. B., Hurwitz J. DNA replication in Escherichia coli made permeable by treatment with high sucrose. Biochem Biophys Res Commun. 1972 Apr 14;47(1):202–211. doi: 10.1016/s0006-291x(72)80029-9. [DOI] [PubMed] [Google Scholar]
  20. Wovcha M. G., Chiu C. S., Tomich P. K., Greenberg G. R. Replicative bacteriophage DNA synthesis in plasmolyzed T4-infected cells: evidence for two independent pathways to DNA. J Virol. 1976 Oct;20(1):142–156. doi: 10.1128/jvi.20.1.142-156.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Wovcha M. G., Tomich P. K., Chiu C. S., Greenberg G. R. Direct participation of dCMP hydroxymethylase in synthesis of bacteriophage T4 DNA. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2196–2200. doi: 10.1073/pnas.70.8.2196. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Yegian C. D., Mueller M., Selzer G., Russo V., Stahl F. W. Properties of the DNA-delay mutants of bacteriophage T4. Virology. 1971 Dec;46(3):900–919. doi: 10.1016/0042-6822(71)90090-0. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES