Roles of topoisomerases in maintaining steady-state DNA supercoiling in Escherichia coli

J Biol Chem. 2000 Mar 17;275(11):8103-13. doi: 10.1074/jbc.275.11.8103.


DNA supercoiling is essential for bacterial cell survival. We demonstrated that DNA topoisomerase IV, acting in concert with topoisomerase I and gyrase, makes an important contribution to the steady-state level of supercoiling in Escherichia coli. Following inhibition of gyrase, topoisomerase IV alone relaxed plasmid DNA to a final supercoiling density (sigma) of -0.015 at an initial rate of 0.8 links min(-1). Topoisomerase I relaxed DNA at a faster rate, 5 links min(-1), but only to a sigma of -0.05. Inhibition of topoisomerase IV in wild-type cells increased supercoiling to approximately the same level as in a mutant lacking topoisomerase I activity (to sigma = -0.08). The role of topoisomerase IV was revealed by two functional assays. Removal of both topoisomerase I and topoisomerase IV caused the DNA to become hyper-negatively supercoiled (sigma = -0.09), greatly stimulating transcription from the supercoiling sensitive leu-500 promoter and increasing the number of supercoils trapped by lambda integrase site-specific recombination.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • DNA Topoisomerase IV
  • DNA Topoisomerases, Type I / metabolism*
  • DNA Topoisomerases, Type II / genetics
  • DNA Topoisomerases, Type II / metabolism*
  • DNA, Bacterial / metabolism*
  • DNA, Superhelical / metabolism*
  • Escherichia coli / enzymology
  • Escherichia coli / genetics*
  • Models, Genetic
  • Mutation
  • Norfloxacin / pharmacology
  • Plasmids / metabolism
  • Recombination, Genetic
  • Topoisomerase II Inhibitors


  • DNA, Bacterial
  • DNA, Superhelical
  • Topoisomerase II Inhibitors
  • DNA Topoisomerase IV
  • DNA Topoisomerases, Type I
  • DNA Topoisomerases, Type II
  • Norfloxacin