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. 1997 Jul 15;25(14):2716–2722. doi: 10.1093/nar/25.14.2716

DNA gyrase can cleave short DNA fragments in the presence of quinolone drugs.

M E Cove 1, A P Tingey 1, A Maxwell 1
PMCID: PMC146802  PMID: 9207016

Abstract

We have analysed the DNA cleavage reaction of DNA gyrase using oligonucleotides annealed to a single-stranded M13 derivative containing a preferred gyrase cleavage site. We find that gyrase can cleave duplexes down to approximately 20 bp in size in the presence of the quinolone drugs ciprofloxacin and oxolinic acid. Ciprofloxacin shows a variation in its site specificity with an apparent preference for G bases adjacent to the cleavage sites, whereas oxolinic acid stimulates cleavage predominantly at the previously determined site. With either drug, cleavage will not occur within 6 bases from the end of a DNA duplex or a nick. We suggest that cleavage site specificity with short DNA duplexes is determined by drug-DNA interactions whereas with longer fragments the positioning effect of the DNA wrap around gyrase prescribes the site of cleavage.

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Selected References

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  1. Ali J. A., Jackson A. P., Howells A. J., Maxwell A. The 43-kilodalton N-terminal fragment of the DNA gyrase B protein hydrolyzes ATP and binds coumarin drugs. Biochemistry. 1993 Mar 16;32(10):2717–2724. doi: 10.1021/bi00061a033. [DOI] [PubMed] [Google Scholar]
  2. Anderson A. H., Sørensen B. S., Christiansen K., Svejstrup J. Q., Lund K., Westergaard O. Studies of the topoisomerase II-mediated cleavage and religation reactions by use of a suicidal double-stranded DNA substrate. J Biol Chem. 1991 May 15;266(14):9203–9210. [PubMed] [Google Scholar]
  3. Bates A. D., Maxwell A. DNA gyrase can supercoil DNA circles as small as 174 base pairs. EMBO J. 1989 Jun;8(6):1861–1866. doi: 10.1002/j.1460-2075.1989.tb03582.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Burgin A. B., Jr, Huizenga B. N., Nash H. A. A novel suicide substrate for DNA topoisomerases and site-specific recombinases. Nucleic Acids Res. 1995 Aug 11;23(15):2973–2979. doi: 10.1093/nar/23.15.2973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Caron P. R., Wang J. C. Appendix. II: Alignment of primary sequences of DNA topoisomerases. Adv Pharmacol. 1994;29B:271–297. doi: 10.1016/s1054-3589(08)61143-6. [DOI] [PubMed] [Google Scholar]
  6. Chen A. Y., Liu L. F. DNA topoisomerases: essential enzymes and lethal targets. Annu Rev Pharmacol Toxicol. 1994;34:191–218. doi: 10.1146/annurev.pa.34.040194.001203. [DOI] [PubMed] [Google Scholar]
  7. Critchlow S. E., Maxwell A. DNA cleavage is not required for the binding of quinolone drugs to the DNA gyrase-DNA complex. Biochemistry. 1996 Jun 11;35(23):7387–7393. doi: 10.1021/bi9603175. [DOI] [PubMed] [Google Scholar]
  8. Dobbs S. T., Cullis P. M., Maxwell A. The cleavage of DNA at phosphorothioate internucleotidic linkages by DNA gyrase. Nucleic Acids Res. 1992 Jul 25;20(14):3567–3573. doi: 10.1093/nar/20.14.3567. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Drlica K., Coughlin S. Inhibitors of DNA gyrase. Pharmacol Ther. 1989;44(1):107–121. doi: 10.1016/0163-7258(89)90093-4. [DOI] [PubMed] [Google Scholar]
  10. Fisher L. M., Barot H. A., Cullen M. E. DNA gyrase complex with DNA: determinants for site-specific DNA breakage. EMBO J. 1986 Jun;5(6):1411–1418. doi: 10.1002/j.1460-2075.1986.tb04375.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fisher L. M., Mizuuchi K., O'Dea M. H., Ohmori H., Gellert M. Site-specific interaction of DNA gyrase with DNA. Proc Natl Acad Sci U S A. 1981 Jul;78(7):4165–4169. doi: 10.1073/pnas.78.7.4165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Froelich-Ammon S. J., Osheroff N. Topoisomerase poisons: harnessing the dark side of enzyme mechanism. J Biol Chem. 1995 Sep 15;270(37):21429–21432. doi: 10.1074/jbc.270.37.21429. [DOI] [PubMed] [Google Scholar]
  13. Gellert M., Fisher L. M., Ohmori H., O'Dea M. H., Mizuuchi K. DNA gyrase: site-specific interactions and transient double-strand breakage of DNA. Cold Spring Harb Symp Quant Biol. 1981;45(Pt 1):391–398. doi: 10.1101/sqb.1981.045.01.053. [DOI] [PubMed] [Google Scholar]
  14. Gellert M., Mizuuchi K., O'Dea M. H., Itoh T., Tomizawa J. I. Nalidixic acid resistance: a second genetic character involved in DNA gyrase activity. Proc Natl Acad Sci U S A. 1977 Nov;74(11):4772–4776. doi: 10.1073/pnas.74.11.4772. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gmünder H., Kuratli K., Keck W. Effect of pyrimido[1,6-a]benzimidazoles, quinolones, and Ca2+ on the DNA gyrase-mediated cleavage reaction. Antimicrob Agents Chemother. 1995 Jan;39(1):163–169. doi: 10.1128/aac.39.1.163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gmünder H., Kuratli K., Keck W. In the presence of subunit A inhibitors DNA gyrase cleaves DNA fragments as short as 20 bp at specific sites. Nucleic Acids Res. 1997 Feb 1;25(3):604–611. doi: 10.1093/nar/25.3.604. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gormley N. A., Orphanides G., Meyer A., Cullis P. M., Maxwell A. The interaction of coumarin antibiotics with fragments of DNA gyrase B protein. Biochemistry. 1996 Apr 16;35(15):5083–5092. doi: 10.1021/bi952888n. [DOI] [PubMed] [Google Scholar]
  18. Hallett P., Grimshaw A. J., Wigley D. B., Maxwell A. Cloning of the DNA gyrase genes under tac promoter control: overproduction of the gyrase A and B proteins. Gene. 1990 Sep 1;93(1):139–142. doi: 10.1016/0378-1119(90)90148-k. [DOI] [PubMed] [Google Scholar]
  19. Higgins N. P., Cozzarelli N. R. The binding of gyrase to DNA: analysis by retention by nitrocellulose filters. Nucleic Acids Res. 1982 Nov 11;10(21):6833–6847. doi: 10.1093/nar/10.21.6833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kampranis S. C., Maxwell A. Conversion of DNA gyrase into a conventional type II topoisomerase. Proc Natl Acad Sci U S A. 1996 Dec 10;93(25):14416–14421. doi: 10.1073/pnas.93.25.14416. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kirkegaard K., Wang J. C. Mapping the topography of DNA wrapped around gyrase by nucleolytic and chemical probing of complexes of unique DNA sequences. Cell. 1981 Mar;23(3):721–729. doi: 10.1016/0092-8674(81)90435-9. [DOI] [PubMed] [Google Scholar]
  22. Lee M. P., Sander M., Hsieh T. Nuclease protection by Drosophila DNA topoisomerase II. Enzyme/DNA contacts at the strong topoisomerase II cleavage sites. J Biol Chem. 1989 Dec 25;264(36):21779–21787. [PubMed] [Google Scholar]
  23. Lewis R. J., Singh O. M., Smith C. V., Skarzynski T., Maxwell A., Wonacott A. J., Wigley D. B. The nature of inhibition of DNA gyrase by the coumarins and the cyclothialidines revealed by X-ray crystallography. EMBO J. 1996 Mar 15;15(6):1412–1420. [PMC free article] [PubMed] [Google Scholar]
  24. Lockshon D., Morris D. R. Sites of reaction of Escherichia coli DNA gyrase on pBR322 in vivo as revealed by oxolinic acid-induced plasmid linearization. J Mol Biol. 1985 Jan 5;181(1):63–74. doi: 10.1016/0022-2836(85)90324-9. [DOI] [PubMed] [Google Scholar]
  25. Lund K., Andersen A. H., Christiansen K., Svejstrup J. Q., Westergaard O. Minimal DNA requirement for topoisomerase II-mediated cleavage in vitro. J Biol Chem. 1990 Aug 15;265(23):13856–13863. [PubMed] [Google Scholar]
  26. Maxwell A. DNA gyrase as a drug target. Trends Microbiol. 1997 Mar;5(3):102–109. doi: 10.1016/S0966-842X(96)10085-8. [DOI] [PubMed] [Google Scholar]
  27. Maxwell A., Gellert M. The DNA dependence of the ATPase activity of DNA gyrase. J Biol Chem. 1984 Dec 10;259(23):14472–14480. [PubMed] [Google Scholar]
  28. Maxwell A. The interaction between coumarin drugs and DNA gyrase. Mol Microbiol. 1993 Aug;9(4):681–686. doi: 10.1111/j.1365-2958.1993.tb01728.x. [DOI] [PubMed] [Google Scholar]
  29. Maxwell A. The molecular basis of quinolone action. J Antimicrob Chemother. 1992 Oct;30(4):409–414. doi: 10.1093/jac/30.4.409. [DOI] [PubMed] [Google Scholar]
  30. Morrison A., Cozzarelli N. R. Contacts between DNA gyrase and its binding site on DNA: features of symmetry and asymmetry revealed by protection from nucleases. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1416–1420. doi: 10.1073/pnas.78.3.1416. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Morrison A., Cozzarelli N. R. Site-specific cleavage of DNA by E. coli DNA gyrase. Cell. 1979 May;17(1):175–184. doi: 10.1016/0092-8674(79)90305-2. [DOI] [PubMed] [Google Scholar]
  32. Orphanides G., Maxwell A. Evidence for a conformational change in the DNA gyrase-DNA complex from hydroxyl radical footprinting. Nucleic Acids Res. 1994 May 11;22(9):1567–1575. doi: 10.1093/nar/22.9.1567. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Pato M. L., Howe M. M., Higgins N. P. A DNA gyrase-binding site at the center of the bacteriophage Mu genome is required for efficient replicative transposition. Proc Natl Acad Sci U S A. 1990 Nov;87(22):8716–8720. doi: 10.1073/pnas.87.22.8716. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Rau D. C., Gellert M., Thoma F., Maxwell A. Structure of the DNA gyrase-DNA complex as revealed by transient electric dichroism. J Mol Biol. 1987 Feb 5;193(3):555–569. doi: 10.1016/0022-2836(87)90266-x. [DOI] [PubMed] [Google Scholar]
  35. Reece R. J., Maxwell A. DNA gyrase: structure and function. Crit Rev Biochem Mol Biol. 1991;26(3-4):335–375. doi: 10.3109/10409239109114072. [DOI] [PubMed] [Google Scholar]
  36. Reece R. J., Maxwell A. Probing the limits of the DNA breakage-reunion domain of the Escherichia coli DNA gyrase A protein. J Biol Chem. 1991 Feb 25;266(6):3540–3546. [PubMed] [Google Scholar]
  37. Reece R. J., Maxwell A. The C-terminal domain of the Escherichia coli DNA gyrase A subunit is a DNA-binding protein. Nucleic Acids Res. 1991 Apr 11;19(7):1399–1405. doi: 10.1093/nar/19.7.1399. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Reece R. J., Maxwell A. Tryptic fragments of the Escherichia coli DNA gyrase A protein. J Biol Chem. 1989 Nov 25;264(33):19648–19653. [PubMed] [Google Scholar]
  39. Rádl S. Structure-activity relationships in DNA gyrase inhibitors. Pharmacol Ther. 1990;48(1):1–17. doi: 10.1016/0163-7258(90)90014-s. [DOI] [PubMed] [Google Scholar]
  40. Schmidt V. K., Sørensen B. S., Sørensen H. V., Alsner J., Westergaard O. Intramolecular and intermolecular DNA ligation mediated by topoisomerase II. J Mol Biol. 1994 Aug 5;241(1):18–25. doi: 10.1006/jmbi.1994.1469. [DOI] [PubMed] [Google Scholar]
  41. Shen L. L., Baranowski J., Pernet A. G. Mechanism of inhibition of DNA gyrase by quinolone antibacterials: specificity and cooperativity of drug binding to DNA. Biochemistry. 1989 May 2;28(9):3879–3885. doi: 10.1021/bi00435a038. [DOI] [PubMed] [Google Scholar]
  42. Sugino A., Cozzarelli N. R. The intrinsic ATPase of DNA gyrase. J Biol Chem. 1980 Jul 10;255(13):6299–6306. [PubMed] [Google Scholar]
  43. Sugino A., Peebles C. L., Kreuzer K. N., Cozzarelli N. R. Mechanism of action of nalidixic acid: purification of Escherichia coli nalA gene product and its relationship to DNA gyrase and a novel nicking-closing enzyme. Proc Natl Acad Sci U S A. 1977 Nov;74(11):4767–4771. doi: 10.1073/pnas.74.11.4767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Svejstrup J. Q., Christiansen K., Andersen A. H., Lund K., Westergaard O. Minimal DNA duplex requirements for topoisomerase I-mediated cleavage in vitro. J Biol Chem. 1990 Jul 25;265(21):12529–12535. [PubMed] [Google Scholar]
  45. Svejstrup J. Q., Christiansen K., Gromova I. I., Andersen A. H., Westergaard O. New technique for uncoupling the cleavage and religation reactions of eukaryotic topoisomerase I. The mode of action of camptothecin at a specific recognition site. J Mol Biol. 1991 Dec 5;222(3):669–678. doi: 10.1016/0022-2836(91)90503-x. [DOI] [PubMed] [Google Scholar]
  46. Sørensen B. S., Sinding J., Andersen A. H., Alsner J., Jensen P. B., Westergaard O. Mode of action of topoisomerase II-targeting agents at a specific DNA sequence. Uncoupling the DNA binding, cleavage and religation events. J Mol Biol. 1992 Dec 5;228(3):778–786. doi: 10.1016/0022-2836(92)90863-f. [DOI] [PubMed] [Google Scholar]
  47. Tamura J. K., Bates A. D., Gellert M. Slow interaction of 5'-adenylyl-beta,gamma-imidodiphosphate with Escherichia coli DNA gyrase. Evidence for cooperativity in nucleotide binding. J Biol Chem. 1992 May 5;267(13):9214–9222. [PubMed] [Google Scholar]
  48. Wahle E., Kornberg A. The partition locus of plasmid pSC101 is a specific binding site for DNA gyrase. EMBO J. 1988 Jun;7(6):1889–1895. doi: 10.1002/j.1460-2075.1988.tb03022.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Wang J. C. DNA topoisomerases. Annu Rev Biochem. 1996;65:635–692. doi: 10.1146/annurev.bi.65.070196.003223. [DOI] [PubMed] [Google Scholar]
  50. Wigley D. B., Davies G. J., Dodson E. J., Maxwell A., Dodson G. Crystal structure of an N-terminal fragment of the DNA gyrase B protein. Nature. 1991 Jun 20;351(6328):624–629. doi: 10.1038/351624a0. [DOI] [PubMed] [Google Scholar]
  51. Willmott C. J., Critchlow S. E., Eperon I. C., Maxwell A. The complex of DNA gyrase and quinolone drugs with DNA forms a barrier to transcription by RNA polymerase. J Mol Biol. 1994 Sep 30;242(4):351–363. doi: 10.1006/jmbi.1994.1586. [DOI] [PubMed] [Google Scholar]
  52. Yang Y., Ames G. F. DNA gyrase binds to the family of prokaryotic repetitive extragenic palindromic sequences. Proc Natl Acad Sci U S A. 1988 Dec;85(23):8850–8854. doi: 10.1073/pnas.85.23.8850. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]

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