PH-dependent salting-out effect in enzyme-catalyzed reaction kinetics

Abstract

The influence of KCl on the second-order rate constant k II = k 2/K s of acetylcholinesterase-catalyzed hydrolysis of butyl acetate is quantitatively described by the linear equation log k II = log k II 0 + Δκc , where c is the concentration and Δκ = κ E − κ X∗ + κ S is the difference between the salting-out coefficients of the initial reagents (E and S) and of the transition state (X ∗. The salting-out parameter Δκ increases with increasing pH (pH range 4.8–6.0) up to the constant value at pH 6.0–8.0, where Δκ = κ S . The pH-dependent Δκ is discussed as an indication of the change in the volume and/or solvation of the enzyme upon the formation of the transition state. The data show that while the pK a1 = 5.6 ± 0.1 and pK a2 = 8.0 ± 0.1 of k II are independent of the KCl concentration, the apparent pK a shift can be observed at high salt concentrations due to a pH-dependent salt effect.