Fig. 1. (A) Purification profile of GyrI. Lane 1, total cell extract after induction; lane 2, sonicate; lane 3, ammonium sulfate pellet; lane 4, pure GyrI after Superdex S200. M, molecular weight marker. The effect of GyrI on supercoiling (B) and relaxation (C) activity. Reactions were carried out with 17.5 nM (supercoiling) and 175 nM (relaxation) DNA gyrase in the presence of 0, 0.17, 0.35, 0.69 and 1.38 µM GyrI (lanes 1–5). R and S represent relaxed and supercoiled pUC18 DNA, respectively. (D) Effect of GyrI on ATPase activity. Reactions were performed with 1.44 µM GyrB in the presence of 0, 0.29, 0.58, 1.17 and 2.34 µM GyrI. ATP (2 mM) was present in all reactions. In addition to GyrB, the +DNA reactions contained 1.4 µM GyrA and 150 µg/ml sonicated salmon sperm DNA. The average of three independent experiments is denoted graphically. All reactions (in B–D) were performed at 37°C for 30 min.