Figure 1.

Effect of residue 225 on the Na⁺ specificity of thrombin, measured as the ratio of the k_cat/K_m values for the hydrolysis of H-D-Phe-Pro-Arg-p-nitroanilide (FPR) in the presence of 200 mM NaCl or choline chloride, 5 mM Tris, 0.1% polyethylene glycol, pH 8.0 at 25°C (6). Binding of Na⁺ to wild type (Tyr) enhances k_cat/K_m nearly 25-fold. A significant enhancement also is observed for Phe, His, Met, Gln, Trp, and Ser, in decreasing order. All other residues show no significant difference between Na⁺ and choline.