Positively supercoiled plasmid DNA is produced by treatment of Escherichia coli with DNA gyrase inhibitors.
A procedure has been developed whereby the relative amounts of the topoisomers of E. coli plasmid can be determined for cells grown under a variety of conditions. Several applications of the procedure are presented. Addition of either novobiocin or oxolinic acid, two inhibitors of DNA gyrase, gives rise to positively supercoiled plasmid. A likely model for the introduction of positive supercoils, involving DNA gyrase itself, is discussed. Oxolinic acid is also shown to induce linearization of plasmid in vivo. Starvation of cells for ATP is shown to cause relaxation of plasmid. The shift of a gyrB temperature-sensitive strain to the restrictive temperature is also shown to cause plasmid relaxation. Finally, it is noted that polyamine starvation of E. coli has no detectable effect on the distribution of topoisomers.
D049933:Aminocoumarins; D003374:Coumarins / Q000494:pharmacology*; D004261:DNA Replication / Q000187:drug effects*; D004278:DNA, Superhelical / Q000235:genetics*; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D009675:Novobiocin / Q000494:pharmacology*; D009693:Nucleic Acid Hybridization; D010093:Oxolinic Acid / Q000494:pharmacology*; D010957:Plasmids / Q000187:drug effects*; D059005:Topoisomerase II Inhibitors
D049933:Aminocoumarins; D003374:Coumarins; D004278:DNA, Superhelical; D059005:Topoisomerase II Inhibitors; D009675:Novobiocin; D010093:Oxolinic Acid; C004628:coumermycin