Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1988 Aug;170(8):3682–3688. doi: 10.1128/jb.170.8.3682-3688.1988

Dominant lethal mutations in the dnaB helicase gene of Salmonella typhimurium.

R Maurer 1, A Wong 1
PMCID: PMC211345  PMID: 2841295

Abstract

A class of dominant lethal mutations in the dnaB (replicative helicase) gene of Salmonella typhimurium is described. The mutated genes, when present on multicopy plasmids, interfered with colony formation by Escherichia coli host strains with a functional chromosomal dnaB gene. The lethal phenotype was expressed specifically in supE (glutamine-inserting) host strains and not in Sup+ strains, because the mutant genes, by design, also possessed an amber mutation derived from a glutamine codon. Mutations located at 11 sites by deletion mapping and DNA sequence analysis varied in the temperature dependence and severity of their lethal effects. None of the mutations complemented a dnaB(Ts) host strain at high temperature (42 degrees C). Therefore, these nonfunctional DnaB proteins must engage some component(s) of the DNA replication machinery and inhibit replication. These mutations are predicted to confer limited, specific defects in either the catalytic activity of DnaB or the ability of DnaB to interact with one of its ligands such as DNA, nucleotide, or another replication protein. The variety of mutant sites and detailed phenotypes represented in this group of mutations may indicate the operation of more than one specific mechanism of lethality.

Full text

PDF
3684

Selected References

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

  1. Arai K., Kornberg A. A general priming system employing only dnaB protein and primase for DNA replication. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4308–4312. doi: 10.1073/pnas.76.9.4308. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. 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]
  3. Arai K., Kornberg A. Mechanism of dnaB protein action. III. Allosteric role of ATP in the alteration of DNA structure by dnaB protein in priming replication. J Biol Chem. 1981 May 25;256(10):5260–5266. [PubMed] [Google Scholar]
  4. Arai K., Kornberg A. Mechanism of dnaB protein action. IV. General priming of DNA replication by dnaB protein and primase compared with RNA polymerase. J Biol Chem. 1981 May 25;256(10):5267–5272. [PubMed] [Google Scholar]
  5. Arai K., Low R., Kobori J., Shlomai J., Kornberg A. Mechanism of dnaB protein action. V. Association of dnaB protein, protein n', and other repriming proteins in the primosome of DNA replication. J Biol Chem. 1981 May 25;256(10):5273–5280. [PubMed] [Google Scholar]
  6. Arai K., McMacken R., Yasuda S., Kornberg A. Purification and properties of Escherichia coli protein i, a prepriming protein in phi X174 DNA replication. J Biol Chem. 1981 May 25;256(10):5281–5286. [PubMed] [Google Scholar]
  7. Arai K., Yasuda S., Kornberg A. Mechanism of dnaB protein action. I. Crystallization and properties of dnaB protein, an essential replication protein in Escherichia coli. J Biol Chem. 1981 May 25;256(10):5247–5252. [PubMed] [Google Scholar]
  8. Backhaus H., Petri J. B. Sequence analysis of a region from the early right operon in phage P22 including the replication genes 18 and 12. Gene. 1984 Dec;32(3):289–303. doi: 10.1016/0378-1119(84)90004-0. [DOI] [PubMed] [Google Scholar]
  9. Baker T. A., Funnell B. E., Kornberg A. Helicase action of dnaB protein during replication from the Escherichia coli chromosomal origin in vitro. J Biol Chem. 1987 May 15;262(14):6877–6885. [PubMed] [Google Scholar]
  10. Baker T. A., Sekimizu K., Funnell B. E., Kornberg A. Extensive unwinding of the plasmid template during staged enzymatic initiation of DNA replication from the origin of the Escherichia coli chromosome. Cell. 1986 Apr 11;45(1):53–64. doi: 10.1016/0092-8674(86)90537-4. [DOI] [PubMed] [Google Scholar]
  11. Cox E. C., Horner D. L. Structure and coding properties of a dominant Escherichia coli mutator gene, mutD. Proc Natl Acad Sci U S A. 1983 Apr;80(8):2295–2299. doi: 10.1073/pnas.80.8.2295. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Degnen G. E., Cox E. C. Conditional mutator gene in Escherichia coli: isolation, mapping, and effector studies. J Bacteriol. 1974 Feb;117(2):477–487. doi: 10.1128/jb.117.2.477-487.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dodson M., Echols H., Wickner S., Alfano C., Mensa-Wilmot K., Gomes B., LeBowitz J., Roberts J. D., McMacken R. Specialized nucleoprotein structures at the origin of replication of bacteriophage lambda: localized unwinding of duplex DNA by a six-protein reaction. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7638–7642. doi: 10.1073/pnas.83.20.7638. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dodson M., Roberts J., McMacken R., Echols H. Specialized nucleoprotein structures at the origin of replication of bacteriophage lambda: complexes with lambda O protein and with lambda O, lambda P, and Escherichia coli DnaB proteins. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4678–4682. doi: 10.1073/pnas.82.14.4678. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Finch P. W., Emmerson P. T. The nucleotide sequence of the uvrD gene of E. coli. Nucleic Acids Res. 1984 Jul 25;12(14):5789–5799. doi: 10.1093/nar/12.14.5789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Günther E., Lanka E., Mikolajczyk M., Schuster H. The dnaB protein of Escherichia coli groPB mutants. J Biol Chem. 1981 Oct 25;256(20):10712–10716. [PubMed] [Google Scholar]
  17. Herskowitz I. Functional inactivation of genes by dominant negative mutations. Nature. 1987 Sep 17;329(6136):219–222. doi: 10.1038/329219a0. [DOI] [PubMed] [Google Scholar]
  18. Lanka E., Geschke B., Schuster H. Escherichia coli dnaB mutant defective in DNA initiation: isolation and properties of the dnaB protein. Proc Natl Acad Sci U S A. 1978 Feb;75(2):799–803. doi: 10.1073/pnas.75.2.799. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lanka E., Schuster H. Replication of bacteriophages in Escherichia coli mutants thermosensitive in DNA synthesis. Mol Gen Genet. 1970;106(3):274–285. doi: 10.1007/BF00340386. [DOI] [PubMed] [Google Scholar]
  20. LeBowitz J. H., McMacken R. The Escherichia coli dnaB replication protein is a DNA helicase. J Biol Chem. 1986 Apr 5;261(10):4738–4748. [PubMed] [Google Scholar]
  21. Marians K. J., Minden J. S., Parada C. Replication of superhelical DNAs in vitro. Prog Nucleic Acid Res Mol Biol. 1986;33:111–140. doi: 10.1016/s0079-6603(08)60021-5. [DOI] [PubMed] [Google Scholar]
  22. Maurer R., Osmond B. C., Botstein D. Genetic analysis of DNA replication in bacteria: dnaB mutations that suppress dnaC mutations and dnaQ mutations that suppress dnaE mutations in Salmonella typhimurium. Genetics. 1984 Sep;108(1):25–38. doi: 10.1093/genetics/108.1.25. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Maurer R., Osmond B. C., Shekhtman E., Wong A., Botstein D. Functional interchangeability of DNA replication genes in Salmonella typhimurium and Escherichia coli demonstrated by a general complementation procedure. Genetics. 1984 Sep;108(1):1–23. doi: 10.1093/genetics/108.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. McMilin K. D., Russo V. E. Maturation and recombination of bacteriophage lambda DNA molecules in the absence of DNA duplication. J Mol Biol. 1972 Jul 14;68(1):49–55. doi: 10.1016/0022-2836(72)90261-6. [DOI] [PubMed] [Google Scholar]
  25. Norrander J., Kempe T., Messing J. Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene. 1983 Dec;26(1):101–106. doi: 10.1016/0378-1119(83)90040-9. [DOI] [PubMed] [Google Scholar]
  26. Rasched I., Oberer E. Ff coliphages: structural and functional relationships. Microbiol Rev. 1986 Dec;50(4):401–427. doi: 10.1128/mr.50.4.401-427.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Reha-Krantz L. J., Hurwitz J. The dnaB gene product of Escherichia coli. I. Purification, homogeneity, and physical properties. J Biol Chem. 1978 Jun 10;253(11):4043–4050. [PubMed] [Google Scholar]
  28. Reha-Krantz L. J., Hurwitz J. The dnaB gene product of Escherichia coli. II. Single stranded DNA-dependent ribonucleoside triphosphatase activity. J Biol Chem. 1978 Jun 10;253(11):4051–4057. [PubMed] [Google Scholar]
  29. Scott J. R. Regulation of plasmid replication. Microbiol Rev. 1984 Mar;48(1):1–23. doi: 10.1016/b978-0-12-048850-6.50006-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. TZAGOLOFF H., PRATT D. THE INITIAL STEPS IN INFECTION WITH COLIPHAGE M13. Virology. 1964 Nov;24:372–380. doi: 10.1016/0042-6822(64)90174-6. [DOI] [PubMed] [Google Scholar]
  31. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  32. Wickner S. DNA-dependent ATPase activity associated with phage P22 gene 12 protein. J Biol Chem. 1984 Nov 25;259(22):14038–14043. [PubMed] [Google Scholar]
  33. Zagursky R. J., Berman M. L. Cloning vectors that yield high levels of single-stranded DNA for rapid DNA sequencing. Gene. 1984 Feb;27(2):183–191. doi: 10.1016/0378-1119(84)90139-2. [DOI] [PubMed] [Google Scholar]

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

RESOURCES