Staphylococcal nuclease digestion of the complex between DNA and DNA gyrase yields a gyrase-DNA core particle composed of a 140 base pair DNA segment and an active gyrase enzyme. The partial specific volume and S20,w of this purified core complex are measured to be 0.70 cm3/g and 14.5 S, respectively, by sedimentation measurements in H2O and D2O media. The molecular weight of the core complex estimated from equilibrium centrifugation is 470 000; the ratio of the translational frictional coefficient to that of the unsolvated equivalent sphere is calculated to be 1.9. Treatment of free gyrase in solution with dimethyl suberimidate gives three cross-linked species of roughly equal amounts that can be identified as alpha 2, alpha 2 beta, and alpha 2 beta 2. When the gyrase core complex is treated with the same cross-linking agent, 70-80% of the protein is converted to the alpha 2 beta 2 species. These results establish that the gyrase-DNA core complex contains a 140 base pair DNA segment and a tetrameric alpha 2 beta 2 protein.