The article describes the purification and characterization of DNA gyrase, an enzyme that converts relaxed closed-circular DNA into negatively supercoiled DNA. The enzyme was purified from Escherichia coli cells and found to require ATP and Mg2+ for its activity, with spermidine stimulating the reaction. DNA gyrase specifically acts on relaxed closed-circular DNA from colicin E1, phage λ, and simian virus 40. The final superhelix density of the DNA can be significantly higher than that found in intracellularly supercoiled DNA. The purification process involved several steps, including fractionation, precipitation, and column chromatography, resulting in a highly active enzyme fraction. The enzyme's mechanism of action is discussed, suggesting that it introduces superhelical turns directly into DNA, rather than through a co-purifying DNA-binding protein and nicking-closing enzyme. The article also explores the potential role of DNA gyrase in DNA replication and its inhibition by novobiocin, indicating its essential function in double-stranded circular DNA replication.The article describes the purification and characterization of DNA gyrase, an enzyme that converts relaxed closed-circular DNA into negatively supercoiled DNA. The enzyme was purified from Escherichia coli cells and found to require ATP and Mg2+ for its activity, with spermidine stimulating the reaction. DNA gyrase specifically acts on relaxed closed-circular DNA from colicin E1, phage λ, and simian virus 40. The final superhelix density of the DNA can be significantly higher than that found in intracellularly supercoiled DNA. The purification process involved several steps, including fractionation, precipitation, and column chromatography, resulting in a highly active enzyme fraction. The enzyme's mechanism of action is discussed, suggesting that it introduces superhelical turns directly into DNA, rather than through a co-purifying DNA-binding protein and nicking-closing enzyme. The article also explores the potential role of DNA gyrase in DNA replication and its inhibition by novobiocin, indicating its essential function in double-stranded circular DNA replication.