MODULATION OF GYRASE-MEDIATED DNA CLEAVAGE AND CELL-KILLING BY ATP

uncoupler of oxidative phosphorylation, 2,4-dinitrophenol, and an acon itase inhibitor, fluoroacetic acid, both of which are known to lower t he cellular ATP pool, protected Escherichia call cells from the bacter icidal actions of gyrase poisons including quinolone antibiotics, nali dixic acid and ciprofloxacin, and the epipodophyllotoxins VP-16 and VM -26. Using purified E. coli DNA gyrase, we examined the effect of ATP on gyrase-mediated DNA cleavage in the presence of these gyrase poison s. ATP was shown to stimulate gyrase-mediated DNA cleavage from HO-to more than 100-fold in the presence of these gyrase poisons, ADP antago nized the stimulatory effect of ATP, consequently, gyrase-mediated DNA cleavage induced by gyrase poisons is modulated by the ATP concentrat ion/ADP concentration ([ATP]/[ADP]) ratio. Coumermycin Al, an inhibito r of the ATPase subunit of DNA gyrase, like ADP, also effectively anta gonized the stimulatory effect of ATP on gyrase-mediated DNA cleavage induced by gyrase poisons. Furthermore, coumermycin Al, like DNP and f luoroacetic acid, also protected cells from the bactericidal action of gyrase poisons, In the aggregate, our results are consistent with the notion that the [ATP]/[ADP] ratio, through its modulatory effect on t he gyrase-mediated DNA cleavage, is an important determinant of cellul ar susceptibility to gyrase poisons.