A library of ciprofloxacin−nuclease conjugates was designed and synthesized to investigate their potential as catalytic antibiotics. The Cu(II) complexes of the new designer compounds (i) showed excellent in vitro hydrolytic and oxidative DNase activity, (ii) showed good antibacterial activity against both Gramnegative and Gram-positive bacteria, and (iii) proved to be highly potent bacterial DNA gyrase inhibitors via a mechanism that involves stabilization of the fluoroquinolone−topoisomerase− DNA ternary complex. Furthermore, the Cu(II) complexes of two of the new designer compounds were shown to fragment supercoiled plasmid DNA into linear DNA in the presence of DNA gyrase, demonstrating a "proof of concept" in vitro. These ciprofloxacin−nuclease conjugates can therefore serve as models with which to develop next-generation, in vivo functioning catalytic antimicrobials.