Skip to main content

Some NLM-NCBI services and products are experiencing heavy traffic, which may affect performance and availability. We apologize for the inconvenience and appreciate your patience. For assistance, please contact our Help Desk at info@ncbi.nlm.nih.gov.

Antimicrobial Agents and Chemotherapy logoLink to Antimicrobial Agents and Chemotherapy
. 1993 Apr;37(4):839–845. doi: 10.1128/aac.37.4.839

Mechanism of action of quinolones against Escherichia coli DNA gyrase.

H Yoshida 1, M Nakamura 1, M Bogaki 1, H Ito 1, T Kojima 1, H Hattori 1, S Nakamura 1
PMCID: PMC187778  PMID: 8388200

Abstract

The mechanism of action of quinolones was investigated by use of various DNA gyrases reconstituted from wild-type and mutant GyrA and GyrB proteins of Escherichia coli. The quinolone sensitivities of the DNA supercoiling activity of the gyrases were generally parallel to the quinolone susceptibilities of strains having the corresponding enzymes and depended on gyrase subunits but not on substrate DNA. [3H]Enoxacin did not bind to gyrase alone or DNA alone but bound to gyrase-DNA complexes when measured by a gel filtration method. There appeared to be two enoxacin binding phases, at low and high enoxacin concentrations, for the wild-type gyrase-DNA and type 2 GyrB (Lys-447 to Glu) mutant gyrase-DNA complexes but only one enoxacin binding phase at the concentrations used for the GyrA (Ser-83 to Leu) mutant gyrase-DNA and type 1 GyrB (Asp-426 to Asn) mutant gyrase-DNA complexes. New enoxacin binding sites appeared in the presence of enoxacin, and the enoxacin binding affinities for the sites, especially at low enoxacin concentrations, near the MICs for the strains having the corresponding gyrases, correlated well with the enoxacin sensitivities of the gyrases and the MICs. From the results obtained, we propose a quinolone pocket model as the mechanism of action of quinolones, in which quinolones exert their action through binding to a gyrase-DNA complex and the quinolone binding affinities for the complex are determined by both GyrA and GyrB subunits in concert.

Full text

PDF
839

Images in this article

Selected References

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

  1. Gellert M. DNA topoisomerases. Annu Rev Biochem. 1981;50:879–910. doi: 10.1146/annurev.bi.50.070181.004311. [DOI] [PubMed] [Google Scholar]
  2. Gellert M., Fisher L. M., O'Dea M. H. DNA gyrase: purification and catalytic properties of a fragment of gyrase B protein. Proc Natl Acad Sci U S A. 1979 Dec;76(12):6289–6293. doi: 10.1073/pnas.76.12.6289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. 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]
  4. 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]
  5. Higgins N. P., Peebles C. L., Sugino A., Cozzarelli N. R. Purification of subunits of Escherichia coli DNA gyrase and reconstitution of enzymatic activity. Proc Natl Acad Sci U S A. 1978 Apr;75(4):1773–1777. doi: 10.1073/pnas.75.4.1773. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Holmes D. S., Quigley M. A rapid boiling method for the preparation of bacterial plasmids. Anal Biochem. 1981 Jun;114(1):193–197. doi: 10.1016/0003-2697(81)90473-5. [DOI] [PubMed] [Google Scholar]
  7. Kaminsky D., Meltzer R. I. Quinolone antibacterial agents. Oxolinic acid and related compounds. J Med Chem. 1968 Jan;11(1):160–163. doi: 10.1021/jm00307a041. [DOI] [PubMed] [Google Scholar]
  8. Klevan L., Wang J. C. Deoxyribonucleic acid gyrase-deoxyribonucleic acid complex containing 140 base pairs of deoxyribonucleic acid and an alpha 2 beta 2 protein core. Biochemistry. 1980 Nov 11;19(23):5229–5234. doi: 10.1021/bi00564a012. [DOI] [PubMed] [Google Scholar]
  9. Kreuzer K. N., Cozzarelli N. R. Escherichia coli mutants thermosensitive for deoxyribonucleic acid gyrase subunit A: effects on deoxyribonucleic acid replication, transcription, and bacteriophage growth. J Bacteriol. 1979 Nov;140(2):424–435. doi: 10.1128/jb.140.2.424-435.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. LESHER G. Y., FROELICH E. J., GRUETT M. D., BAILEY J. H., BRUNDAGE R. P. 1,8-NAPHTHYRIDINE DERIVATIVES. A NEW CLASS OF CHEMOTHERAPEUTIC AGENTS. J Med Pharm Chem. 1962 Sep;91:1063–1065. doi: 10.1021/jm01240a021. [DOI] [PubMed] [Google Scholar]
  11. Matsumoto J., Miyamoto T., Minamida A., Nishimura Y., Egawa H., Nishimura H. Pyridonecarboxylic acids as antibacterial agents. 2. Synthesis and structure-activity relationships of 1,6,7-trisubstituted 1,4-dihydro-4-oxo-1,8-naphthyridine-3-carboxylic acids, including enoxacin, a new antibacterial agent. J Med Chem. 1984 Mar;27(3):292–301. doi: 10.1021/jm00369a011. [DOI] [PubMed] [Google Scholar]
  12. Miyamoto T., Matsumoto J., Chiba K., Egawa H., Shibamori K., Minamida A., Nishimura Y., Okada H., Kataoka M., Fujita M. Synthesis and structure-activity relationships of 5-substituted 6,8-difluoroquinolones, including sparfloxacin, a new quinolone antibacterial agent with improved potency. J Med Chem. 1990 Jun;33(6):1645–1656. doi: 10.1021/jm00168a018. [DOI] [PubMed] [Google Scholar]
  13. Mizuuchi K., Mizuuchi M., O'Dea M. H., Gellert M. Cloning and simplified purification of Escherichia coli DNA gyrase A and B proteins. J Biol Chem. 1984 Jul 25;259(14):9199–9201. [PubMed] [Google Scholar]
  14. Mizuuchi K., O'Dea M. H., Gellert M. DNA gyrase: subunit structure and ATPase activity of the purified enzyme. Proc Natl Acad Sci U S A. 1978 Dec;75(12):5960–5963. doi: 10.1073/pnas.75.12.5960. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Nakamura S., Nakamura M., Kojima T., Yoshida H. gyrA and gyrB mutations in quinolone-resistant strains of Escherichia coli. Antimicrob Agents Chemother. 1989 Feb;33(2):254–255. doi: 10.1128/aac.33.2.254. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Sato K., Inoue Y., Fujii T., Aoyama H., Inoue M., Mitsuhashi S. Purification and properties of DNA gyrase from a fluoroquinolone-resistant strain of Escherichia coli. Antimicrob Agents Chemother. 1986 Nov;30(5):777–780. doi: 10.1128/aac.30.5.777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Shen L. L., Kohlbrenner W. E., Weigl D., Baranowski J. Mechanism of quinolone inhibition of DNA gyrase. Appearance of unique norfloxacin binding sites in enzyme-DNA complexes. J Biol Chem. 1989 Feb 15;264(5):2973–2978. [PubMed] [Google Scholar]
  18. Shen L. L., Mitscher L. A., Sharma P. N., O'Donnell T. J., Chu D. W., Cooper C. S., Rosen T., Pernet A. G. Mechanism of inhibition of DNA gyrase by quinolone antibacterials: a cooperative drug--DNA binding model. Biochemistry. 1989 May 2;28(9):3886–3894. doi: 10.1021/bi00435a039. [DOI] [PubMed] [Google Scholar]
  19. Shen L. L., Pernet A. G. Mechanism of inhibition of DNA gyrase by analogues of nalidixic acid: the target of the drugs is DNA. Proc Natl Acad Sci U S A. 1985 Jan;82(2):307–311. doi: 10.1073/pnas.82.2.307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. 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]
  21. Walton L., Elwell L. P. In vitro cleavable-complex assay to monitor antimicrobial potency of quinolones. Antimicrob Agents Chemother. 1988 Jul;32(7):1086–1089. doi: 10.1128/aac.32.7.1086. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Wang J. C. DNA topoisomerases. Annu Rev Biochem. 1985;54:665–697. doi: 10.1146/annurev.bi.54.070185.003313. [DOI] [PubMed] [Google Scholar]
  23. Yamagishi J., Yoshida H., Yamayoshi M., Nakamura S. Nalidixic acid-resistant mutations of the gyrB gene of Escherichia coli. Mol Gen Genet. 1986 Sep;204(3):367–373. doi: 10.1007/BF00331012. [DOI] [PubMed] [Google Scholar]
  24. Yoshida H., Bogaki M., Nakamura M., Nakamura S. Quinolone resistance-determining region in the DNA gyrase gyrA gene of Escherichia coli. Antimicrob Agents Chemother. 1990 Jun;34(6):1271–1272. doi: 10.1128/aac.34.6.1271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Yoshida H., Bogaki M., Nakamura M., Yamanaka L. M., Nakamura S. Quinolone resistance-determining region in the DNA gyrase gyrB gene of Escherichia coli. Antimicrob Agents Chemother. 1991 Aug;35(8):1647–1650. doi: 10.1128/aac.35.8.1647. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Yoshida H., Kojima T., Yamagishi J., Nakamura S. Quinolone-resistant mutations of the gyrA gene of Escherichia coli. Mol Gen Genet. 1988 Jan;211(1):1–7. doi: 10.1007/BF00338386. [DOI] [PubMed] [Google Scholar]

Articles from Antimicrobial Agents and Chemotherapy are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES