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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1983 Apr;80(8):2189–2192. doi: 10.1073/pnas.80.8.2189

Mutator strains of Escherichia coli, mutD and dnaQ, with defective exonucleolytic editing by DNA polymerase III holoenzyme.

H Echols, C Lu, P M Burgers
PMCID: PMC393783  PMID: 6340117

Abstract

The closely linked mutD and dnaQ mutations confer a vastly increased mutation rate on Escherichia coli and thus might define a gene with a central role in the fidelity of DNA replication. To look for the biochemical function of the mutD gene product, we have measured the 3' leads to 5' exonucleolytic editing activity of polymerase III holoenzyme from mutD5 and dnaQ49 mutants. The editing activities of the mutant enzymes are defective compared to wild type, as judged by two assays: (i) decreased excision of a terminal mispaired base from a copolymer substrate and (ii) turnover of dTTP to dTMP during replication with a phage G4 DNA template. Thus, the mutD (dnaQ) gene product is likely to control the editing (proofreading) capacity of polymerase III holoenzyme.

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Selected References

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

  1. Brutlag D., Kornberg A. Enzymatic synthesis of deoxyribonucleic acid. 36. A proofreading function for the 3' leads to 5' exonuclease activity in deoxyribonucleic acid polymerases. J Biol Chem. 1972 Jan 10;247(1):241–248. [PubMed] [Google Scholar]
  2. Clayton L. K., Goodman M. F., Branscomb E. W., Galas D. J. Error induction and correction by mutant and wild type T4 DNA polymerases. Kinetic error discrimination mechanisms. J Biol Chem. 1979 Mar 25;254(6):1902–1912. [PubMed] [Google Scholar]
  3. Cox E. C., Horner D. L. Dominant mutators in Escherichia coli. Genetics. 1982 Jan;100(1):7–18. doi: 10.1093/genetics/100.1.7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. 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]
  5. Drake J. W., Allen E. F., Forsberg S. A., Preparata R. M., Greening E. O. Genetic control of mutation rates in bacteriophageT4. Nature. 1969 Mar 22;221(5186):1128–1132. [PubMed] [Google Scholar]
  6. Drake J. W., Allen E. F., Forsberg S. A., Preparata R. M., Greening E. O. Genetic control of mutation rates in bacteriophageT4. Nature. 1969 Mar 22;221(5186):1128–1132. [PubMed] [Google Scholar]
  7. Echols H. Mutation rate: some biological and biochemical considerations. Biochimie. 1982 Aug-Sep;64(8-9):571–575. doi: 10.1016/s0300-9084(82)80089-8. [DOI] [PubMed] [Google Scholar]
  8. Echols H. SOS functions, cancer and inducible evolution. Cell. 1981 Jul;25(1):1–2. doi: 10.1016/0092-8674(81)90223-3. [DOI] [PubMed] [Google Scholar]
  9. Eisenberg S., Kornberg A. Purification and characterization of phiX174 gene A protein. A multifunctional enzyme of duplex DNA replication. J Biol Chem. 1979 Jun 25;254(12):5328–5332. [PubMed] [Google Scholar]
  10. Fersht A. R., Knill-Jones J. W., Tsui W. C. Kinetic basis of spontaneous mutation. Misinsertion frequencies, proofreading specificities and cost of proofreading by DNA polymerases of Escherichia coli. J Mol Biol. 1982 Mar 25;156(1):37–51. doi: 10.1016/0022-2836(82)90457-0. [DOI] [PubMed] [Google Scholar]
  11. Fowler R. G., Degnen G. E., Cox E. C. Mutational specificity of a conditional Escherichia coli mutator, mutD5. Mol Gen Genet. 1974;133(3):179–191. doi: 10.1007/BF00267667. [DOI] [PubMed] [Google Scholar]
  12. Hershfield M. S., Nossal N. G. Hydrolysis of template and newly synthesized deoxyribonucleic acid by the 3' to 5' exonuclease activity of the T4 deoxyribonucleic acid polymerase. J Biol Chem. 1972 Jun 10;247(11):3393–3404. [PubMed] [Google Scholar]
  13. Horiuchi T., Maki H., Maruyama M., Sekiguchi M. Identification of the dnaQ gene product and location of the structural gene for RNase H of Escherichia coli by cloning of the genes. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3770–3774. doi: 10.1073/pnas.78.6.3770. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Horiuchi T., Maki H., Sekiguchi M. A new conditional lethal mutator (dnaQ49) in Escherichia coli K12. Mol Gen Genet. 1978 Jul 25;163(3):277–283. doi: 10.1007/BF00271956. [DOI] [PubMed] [Google Scholar]
  15. Horiuchi T., Maki H., Sekiguchi M. Conditional lethality of Escherichia coli strains carrying dnaE and dnaQ mutations. Mol Gen Genet. 1981;181(1):24–28. doi: 10.1007/BF00339000. [DOI] [PubMed] [Google Scholar]
  16. Ichikawa-Ryo H., Kondo S. Indirect mutagenesis in phage lambda by ultraviolet preirradiation of host bacteria. J Mol Biol. 1975 Sep 5;97(1):77–92. doi: 10.1016/s0022-2836(75)80023-4. [DOI] [PubMed] [Google Scholar]
  17. Johanson K. O., McHenry C. S. Purification and characterization of the beta subunit of the DNA polymerase III holoenzyme of Escherichia coli. J Biol Chem. 1980 Nov 25;255(22):10984–10990. [PubMed] [Google Scholar]
  18. Loeb L. A., Kunkel T. A. Fidelity of DNA synthesis. Annu Rev Biochem. 1982;51:429–457. doi: 10.1146/annurev.bi.51.070182.002241. [DOI] [PubMed] [Google Scholar]
  19. McHenry C., Kornberg A. DNA polymerase III holoenzyme of Escherichia coli. Purification and resolution into subunits. J Biol Chem. 1977 Sep 25;252(18):6478–6484. [PubMed] [Google Scholar]
  20. Mount D. W. A mutant of Escherichia coli showing constitutive expression of the lysogenic induction and error-prone DNA repair pathways. Proc Natl Acad Sci U S A. 1977 Jan;74(1):300–304. doi: 10.1073/pnas.74.1.300. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Muzyczka N., Poland R. L., Bessman M. J. Studies on the biochemical basis of spontaneous mutation. I. A comparison of the deoxyribonucleic acid polymerases of mutator, antimutator, and wild type strains of bacteriophage T4. J Biol Chem. 1972 Nov 25;247(22):7116–7122. [PubMed] [Google Scholar]
  22. Rowen L., Kornberg A. Primase, the dnaG protein of Escherichia coli. An enzyme which starts DNA chains. J Biol Chem. 1978 Feb 10;253(3):758–764. [PubMed] [Google Scholar]
  23. Spanos A., Sedgwick S. G., Yarranton G. T., Hübscher U., Banks G. R. Detection of the catalytic activities of DNA polymerases and their associated exonucleases following SDS-polyacrylamide gel electrophoresis. Nucleic Acids Res. 1981 Apr 24;9(8):1825–1839. doi: 10.1093/nar/9.8.1825. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Topal M. D., Fresco J. R. Complementary base pairing and the origin of substitution mutations. Nature. 1976 Sep 23;263(5575):285–289. doi: 10.1038/263285a0. [DOI] [PubMed] [Google Scholar]
  25. Weiner J. H., Bertsch L. L., Kornberg A. The deoxyribonucleic acid unwinding protein of Escherichia coli. Properties and functions in replication. J Biol Chem. 1975 Mar 25;250(6):1972–1980. [PubMed] [Google Scholar]
  26. Witkin E. M., Wermundsen I. E. Targeted and untargeted mutagenesis by various inducers of SOS functions in Escherichia coli. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 2):881–886. doi: 10.1101/sqb.1979.043.01.095. [DOI] [PubMed] [Google Scholar]
  27. Wold M. S., McMacken R. Regulation of expression of the Escherichia coli dnaG gene and amplification of the dnaG primase. Proc Natl Acad Sci U S A. 1982 Aug;79(16):4907–4911. doi: 10.1073/pnas.79.16.4907. [DOI] [PMC free article] [PubMed] [Google Scholar]

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