Richard M. Burger

Bacterial DNA supercoiling and [ATP][ADP]: Changes associated with a transition to anaerobic growth

Shifting Escherichia coli from aerobic to anaerobic growth caused changes in the ratio of [ATP]/[ADP] and in negative supercoiling of chromosomal and plasmid DNA. Shortly after lowering oxygen tension, both [ATP]/[ADP] and supercoiling transiently decreased. Under conditions of exponential anaerobic growth, both were higher than under aerobic conditions. These correlations may reflect an effect of [ATP]/[ADP] on DNA gyrase, since in vitro [ATP]/[ADP] influences the level of plasmid supercoiling attained when gyrase is either introducing or removing supercoils. When the supercoiling activity of gyrase was perturbed by a mutation in gyrB, a shift to anaerobic conditions resulted in plasmid supercoil relaxation similar to that seen with wild-type. However, the low level of supercoiling in the mutant persisted during a time when supercoiling in wild-type recovered and then exceeded aerobic levels. Thus, changes in oxygen tension can alter DNA supercoiling through an effect on gyrase, and correlations exist between changes in supercoiling and changes in the intracellular ratio of [ATP]/[ADP].

Nature of Activated Bleomycin

Activated bleomycin is the drug species seen to be kinetically competent to initiate DNA degradation in the reaction of bleomycin, Fe(II), and O2. It also forms in reactions of bleomycin with Fe(III) and peroxide, or bleomycin with superoxide and either Fe(III) or Fe(II). Efforts to characterize this transient species proceeded by kinetic and spectroscopic strategies. Activated bleomycin now appears to be a drug-ferric-peroxide complex, but this may not be the proximate active drug species. Assuming that activated bleomycin peroxide cleavage yields a reactive product analogous to peroxidase compound I explains many characteristics of bleomycin-mediated DNA degradation reactions. DNA degradation is responsible for the cytotoxic and antitumor activities of this clinically useful antibiotic.

Killing acanthamoebae with polyaminopropyl biguanide: quantitation and kinetics.

The two Acanthamoeba species most often implicated in corneal keratitis, A. castellanii and A. polyphaga, were exposed as cysts to polyaminopropyl biguanide (PAPB), a commonly used antimicrobial agent. Killing of amoeba cysts was rapid and extensive, with fewer than 2% of either species surviving 30 s of exposure to > or = 45 ppm of PAPB. Killing kinetics were biphasic, and further exposures of 15 min to 1 h killed greater than 90% of those surviving initial killing. This potency of PAPB, together with its low toxicity to humans when ingested or applied topically, underscores the potential of PAPB as an antiamoebic agent.

Superoxide protects Escherichia coli from bleomycin mediated lethality.

Superoxide and its products, especially hydroxyl radical, were recently proposed to be instrumental in cell death following treatment with a wide range of antimicrobials. Surprisingly, bleomycin lethality to Escherichia coli was ameliorated by a genetic deficiency of superoxide dismutase or by furnishing the superoxide generator plumbagin. Rescue by plumbagin was similar in strains containing or lacking recA or with inactive, inducible, or constitutive soxRS regulons. Thus, superoxide interferes with bleomycin cytotoxicity in ways not readily explained by genetic pathways expected to protect from oxidative damage.