Effect of Sodium Chloride and pH on the Rate of Neutralization of Cypridina Luciferase by Specific Antibody

Abstract

Summary The effect of sodium chloride concentration and pH on the kinetics of the neutralization of Cypridina luciferase by antiluciferase γ-globulin has been investigated. Determination of the rate of neutralization in the presence of excess γ-globulin showed that approximately 60% of the initial activity was neutralized according to pseudo-first order kinetics. The rate of neutralization increased on lowering the sodium chloride concentration and decreased on increasing the salt concentration. The rate also increased in going from pH 7.0 to 5.4 at low salt concentrations, but was insensitive to variation in hydrogen ion concentration at high salt concentrations. At low ionic strengths, the observed results conformed to the predictions of the Brönsted theory of the primary salt effect on reaction kinetics. Consequently, the rate changes observed in the presence of sodium and chloride ions were considered as being due to their effect on the “activity” of an intermediate “activated complex.” Alternatively, the effect of salt and hydrogen ion concentration on reaction rate was also considered in terms of a modified diffuse double layer affecting the collision frequency of the molecules. Determination of the rate constants at varying hydrogen ion concentrations, and subsequent analysis of the data in terms of an interaction between a single ionic species of luciferase and antibody, indicated the involvement of active groups possessing pK1 = pK2 = pK3 = 5 and pK4 ≧ 8.