Figure 4.

pH dependence of the rate of substrate cleavage. Single-turnover reactions were carried out at 25°C, in the presence of either 3 mM MgCl₂ (open circles) or 30 mM MgCl₂ (closed circles). The buffer used (50 mM Bis-tris propane–HCl) could cover the entire pH range 6.4–9.4, and its ionic strength was kept constant at 0.2 M by adding the appropriate amount of NaCl at each pH value. The solid lines represent linear least-squares fits of the data up to pH 8.5, and have slopes of 0.98 ± 0.1 (3 mM MgCl₂) and 0.95 ± 0.1 (30 mM MgCl₂). Nearly identical dependencies, with slopes of ∼1.0 in the pH range 6.5–8.5, were observed for reactions carried out in 3 or 30 mM Ca²⁺ (data not shown).