Mechanism of quinolone action. A drug-induced structural perturbation of the DNA precedes strand cleavage by topoisomerase IV.
Quinolones are potent broad spectrum antibacterial drugs that target the bacterial type II DNA topoisomerases. Their cytotoxicity derives from their ability to shift the cleavage-religation equilibrium required for topoisomerase action toward cleavage, thereby effectively trapping the enzyme on the DNA. It has been proposed that these drugs act by binding to the enzyme-DNA complex. Using catalytically inactive and quinolone-resistant mutant topoisomerase IV proteins, nitrocellulose filter DNA binding assays, and KMnO4 probing of drug-DNA and drug-DNA-enzyme complexes, we show: (i) that norfloxacin binding to DNA induces a structural alteration, which probably corresponds to an unwinding of the helix, that is exacerbated by binding of the topoisomerase and by binding of the drug to the enzyme and (ii) that formation of this structural perturbation in the DNA precedes DNA cleavage by the topoisomerase in the ternary complex. We conclude that cleavage of the DNA and the resultant opening of the DNA gate during topoisomerization requires the induction of strain in the DNA that is bound to the enzyme. We suggest that quinolones may act to accelerate the rate of DNA cleavage by stimulating acquisition of this structural perturbation in the ternary complex.
Marians
K J
KJ
Molecular Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Hiasa
H
H
D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000187:drug effects*; D027101:DNA Topoisomerase IV; D004250:DNA Topoisomerases, Type II / Q000235:genetics / Q000378:metabolism*; D004278:DNA, Superhelical / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D016297:Mutagenesis, Site-Directed; D009643:Norfloxacin / Q000494:pharmacology*; D009690:Nucleic Acid Conformation / Q000187:drug effects; D011196:Potassium Permanganate / Q000378:metabolism; D013941:Thymine / Q000378:metabolism
D004278:DNA, Superhelical; D004791:Enzyme Inhibitors; D011196:Potassium Permanganate; D004247:DNA; D027101:DNA Topoisomerase IV; D004250:DNA Topoisomerases, Type II; D009643:Norfloxacin; D013941:Thymine