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Fluoroquinolones inhibit preferentially Streptococcus pneumoniae DNA topoisomerase IV than DNA gyrase native proteins

E Fernandez-Moreira1, D Balas, I Gonzalez

  • 1Unidad de Genética Bacteriana, Consejo Superior de Investigaciones Científicas, Centro Nacional de Biologia Fundamental, Instituto de Salud Carlos III, Majadahonda, Madrid, Spain.

Microbial Drug Resistance (Larchmont, N.Y.)
|March 29, 2001
PubMed

Insights

This study purified DNA topoisomerase IV and DNA gyrase from Streptococcus pneumoniae. Fluoroquinolones showed greater inhibition of topoisomerase IV, suggesting it

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Microbiology

Background:

  • DNA topoisomerase IV and DNA gyrase are essential enzymes in bacteria.
  • Fluoroquinolones are a class of antibiotics that target these enzymes.

Purpose of the Study:

  • To clone, overproduce, and purify subunits of Streptococcus pneumoniae DNA topoisomerase IV and DNA gyrase.
  • To reconstitute the enzymatic activities of both enzymes.
  • To compare the inhibitory effects of four fluoroquinolones on these enzymes.

Main Methods:

  • Cloning and overproduction of parC, parE, gyrA, and gyrB genes in Escherichia coli.
  • Purification of enzyme subunits using DEAE-sepharose, heparin-agarose, and hydroxylapatite chromatography.
  • Reconstitution of DNA topoisomerase IV and DNA gyrase activities.
  • Assay of fluoroquinolone inhibition on DNA decatenation (topoisomerase IV) and supercoiling (gyrase).

Main Results:

  • Reconstituted DNA topoisomerase IV exhibited efficient ATP-dependent DNA decatenation.
  • Reconstituted DNA gyrase showed ATP-dependent supercoiling activity.
  • All tested fluoroquinolones were more potent inhibitors of topoisomerase IV than DNA gyrase.
  • A positive correlation was observed between inhibitory activity against topoisomerase IV decatenation and DNA gyrase supercoiling.
  • Fluoroquinolone efficacy ranking: clinafloxacin > trovafloxacin > sparfloxacin > ciprofloxacin.

Conclusions:

  • The findings indicate that fluoroquinolones primarily target DNA topoisomerase IV in S. pneumoniae.
  • DNA gyrase appears to be a secondary target for these antibiotics.
  • Results align with existing genetic data on fluoroquinolone mechanisms of action.

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