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. 1982 May;79(9):2832–2835. doi: 10.1073/pnas.79.9.2832

Escherichia coli single-strand DNA binding protein from wild type and lexC113 mutant affects in vitro proteolytic cleavage of phage lambda repressor.

J Resnick, R Sussman
PMCID: PMC346300  PMID: 6211675

Abstract

In Escherichia coli, the single-strand DNA-binding protein (SSB) is required for DNA replication. A mutation of the ssb gene, lexC113, imparts to the cells UV sensitivity and inability to induce lambda prophage and to amplify recA protein, indicating participation of SSB in DNA repair and viral induction processes. We report the effect of purified SSB, isolated from wild-type and lexC113 strains, on the recA-mediated proteolysis of lambda repressor in vitro. (i) These proteins abolished the inhibition produced by excess single-strand DNA and (ii) in the presence of the binding proteins, the apparent stoichiometry--1 monomer of recA to 6 nucleotides of single-strand DNA [Craig, N. L. & Roberts, J. W. (1980) Nature (London) 283, 26-30] was no longer observed. (iii) At the optimal concentration--1 protein monomer to 8 nucleotides--they increased the rate and extent of repressor cleavage at all single-strand DNA concentrations, including that observed at the apparent optimal DNA concentration. (iv) At binding protein/nucleotide ratios greater than or equal to 1:3, SSB from lexC113 inhibited repressor cleavage while that from wild type did not. (v) These results are consistent with the notion that SSB is probably involved in the induction of prophages in vivo.

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Selected References

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

  1. Baluch J., Chase J. W., Sussman R. Synthesis of recA protein and induction of bacteriophage lambda in single-strand deoxyribonucleic acid-binding protein mutants of Escherichia coli. J Bacteriol. 1980 Nov;144(2):489–498. doi: 10.1128/jb.144.2.489-498.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brooks K., Clark A. J. Behavior of lambda bacteriophage in a recombination deficienct strain of Escherichia coli. J Virol. 1967 Apr;1(2):283–293. doi: 10.1128/jvi.1.2.283-293.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Chase J. W., Whittier R. F., Auerbach J., Sancar A., Rupp W. D. Amplification of single-strand DNA binding protein in Escherichia coli. Nucleic Acids Res. 1980 Jul 25;8(14):3215–3227. doi: 10.1093/nar/8.14.3215. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Clewell D. B., Helinski D. R. Properties of a supercoiled deoxyribonucleic acid-protein relaxation complex and strand specificity of the relaxation event. Biochemistry. 1970 Oct 27;9(22):4428–4440. doi: 10.1021/bi00824a026. [DOI] [PubMed] [Google Scholar]
  5. Craig N. L., Roberts J. W. E. coli recA protein-directed cleavage of phage lambda repressor requires polynucleotide. Nature. 1980 Jan 3;283(5742):26–30. doi: 10.1038/283026a0. [DOI] [PubMed] [Google Scholar]
  6. Eisenberg S., Griffith J., Kornberg A. phiX174 cistron A protein is a multifunctional enzyme in DNA replication. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3198–3202. doi: 10.1073/pnas.74.8.3198. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Glassberg J., Meyer R. R., Kornberg A. Mutant single-strand binding protein of Escherichia coli: genetic and physiological characterization. J Bacteriol. 1979 Oct;140(1):14–19. doi: 10.1128/jb.140.1.14-19.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Greenberg J., Berends L. J., Donch J., Green M. H. exrB: a malB-linked gene in Escherichia coli B involved in sensitivity to radiation and filament formation. Genet Res. 1974 Apr;23(2):175–184. doi: 10.1017/s0016672300014798. [DOI] [PubMed] [Google Scholar]
  9. Gudas L. J., Pardee A. B. DNA synthesis inhibition and the induction of protein X in Escherichia coli. J Mol Biol. 1976 Mar 15;101(4):459–477. doi: 10.1016/0022-2836(76)90240-0. [DOI] [PubMed] [Google Scholar]
  10. Levine A., Bailone A., Devoret R. Cellular levels of the prophage lambda and 434 repressors. J Mol Biol. 1979 Jul 5;131(3):655–661. doi: 10.1016/0022-2836(79)90014-7. [DOI] [PubMed] [Google Scholar]
  11. Little J. W., Edmiston S. H., Pacelli L. Z., Mount D. W. Cleavage of the Escherichia coli lexA protein by the recA protease. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3225–3229. doi: 10.1073/pnas.77.6.3225. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. McEntee K., Weinstock G. M., Lehman I. R. Initiation of general recombination catalyzed in vitro by the recA protein of Escherichia coli. Proc Natl Acad Sci U S A. 1979 Jun;76(6):2615–2619. doi: 10.1073/pnas.76.6.2615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. McEntee K., Weinstock G. M., Lehman I. R. recA protein-catalyzed strand assimilation: stimulation by Escherichia coli single-stranded DNA-binding protein. Proc Natl Acad Sci U S A. 1980 Feb;77(2):857–861. doi: 10.1073/pnas.77.2.857. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Meyer R. R., Glassberg J., Scott J. V., Kornberg A. A temperature-sensitive single-stranded DNA-binding protein from Escherichia coli. J Biol Chem. 1980 Apr 10;255(7):2897–2901. [PubMed] [Google Scholar]
  15. Molineux I. J., Friedman S., Gefter M. L. Purification and properties of the Escherichia coli deoxyribonucleic acid-unwinding protein. Effects on deoxyribonucleic acid synthesis in vitro. J Biol Chem. 1974 Oct 10;249(19):6090–6098. [PubMed] [Google Scholar]
  16. Ogawa T., Wabiko H., Tsurimoto T., Horii T., Masukata H., Ogawa H. Characteristics of purified recA protein and the regulation of its synthesis in vivo. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 2):909–915. doi: 10.1101/sqb.1979.043.01.099. [DOI] [PubMed] [Google Scholar]
  17. Phizicky E. M., Roberts J. W. Induction of SOS functions: regulation of proteolytic activity of E. coli RecA protein by interaction with DNA and nucleoside triphosphate. Cell. 1981 Jul;25(1):259–267. doi: 10.1016/0092-8674(81)90251-8. [DOI] [PubMed] [Google Scholar]
  18. Phizicky E. M., Roberts J. W. Kinetics of RecA protein-directed inactivation of repressors of phage lambda and phage P22. J Mol Biol. 1980 May 25;139(3):319–328. doi: 10.1016/0022-2836(80)90133-3. [DOI] [PubMed] [Google Scholar]
  19. Roberts J. W., Roberts C. W., Craig N. L., Phizicky E. M. Activity of the Escherichia coli recA-gene product. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 2):917–920. doi: 10.1101/sqb.1979.043.01.100. [DOI] [PubMed] [Google Scholar]
  20. Roberts J. W., Roberts C. W. Proteolytic cleavage of bacteriophage lambda repressor in induction. Proc Natl Acad Sci U S A. 1975 Jan;72(1):147–151. doi: 10.1073/pnas.72.1.147. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Sevastopoulos C. G., Wehr C. T., Glaser D. A. Large-scale automated isolation of Escherichia coli mutants with thermosensitive DNA replication. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3485–3489. doi: 10.1073/pnas.74.8.3485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Shibata T., Cunningham R. P., DasGupta C., Radding C. M. Homologous pairing in genetic recombination: complexes of recA protein and DNA. Proc Natl Acad Sci U S A. 1979 Oct;76(10):5100–5104. doi: 10.1073/pnas.76.10.5100. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Shibata T., DasGupta C., Cunningham R. P., Radding C. M. Homologous pairing in genetic recombination: formation of D loops by combined action of recA protein and a helix-destabilizing protein. Proc Natl Acad Sci U S A. 1980 May;77(5):2606–2610. doi: 10.1073/pnas.77.5.2606. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Shibata T., DasGupta C., Cunningham R. P., Radding C. M. Purified Escherichia coli recA protein catalyzes homologous pairing of superhelical DNA and single-stranded fragments. Proc Natl Acad Sci U S A. 1979 Apr;76(4):1638–1642. doi: 10.1073/pnas.76.4.1638. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Sussman R., Resnick J., Calame K., Baluch J. Interaction of bacteriophage lambda repressor with nonoperator DNA containing single-strand gaps. Proc Natl Acad Sci U S A. 1978 Dec;75(12):5817–5821. doi: 10.1073/pnas.75.12.5817. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Vales L. D., Chase J. W., Murphy J. B. Effect of ssbA1 and lexC113 mutations on lambda prophage induction, bacteriophage growth, and cell survival. J Bacteriol. 1980 Aug;143(2):887–896. doi: 10.1128/jb.143.2.887-896.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Weinstock G. M., McEntee K., Lehman I. R. ATP-dependent renaturation of DNA catalyzed by the recA protein of Escherichia coli. Proc Natl Acad Sci U S A. 1979 Jan;76(1):126–130. doi: 10.1073/pnas.76.1.126. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Weinstock G. M., McEntee K. RecA protein-dependent proteolysis of bacteriophage lambda repressor Characterization of the reaction and stimulation by DNA-binding proteins. J Biol Chem. 1981 Nov 10;256(21):10883–10888. [PubMed] [Google Scholar]
  29. West S. C., Cassuto E., Howard-Flanders P. Homologous pairing can occur before DNA strand separation in general genetic recombination. Nature. 1981 Mar 5;290(5801):29–33. doi: 10.1038/290029a0. [DOI] [PubMed] [Google Scholar]

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