DNA topoisomerases are essential to the cell for the regulation of DNA supercoiling levels and for chromosome decatenation. The proposed mechanisms for these reactions are essentially the same, except that a change in supercoiling is due to an intramolecular event, while decatenation requires an intermolecular event. The characterized bacterial topoisomerases appear capable of both types of reaction in vitro. Four DNA topoisomerases have been identified in Escherichia coli. Topoisomerase I, gyrase, and topoisomerase IV normally appear to have distinct essential functions within the cell. Gyrase and topoisomerase I are responsible for the regulation of DNA supercoiling. Both gyrase and topoisomerase IV are necessary for chromosomal decatenation. Multiple topoisomerases with distinct functions may give the cell more precise control over DNA topology by allowing tighter regulation of the principal enzymatic activities of these different proteins.