Kinetic studies of a (Na++ K++ Mg2+)ATPase in sugar beet roots. III. A proposed model for the (Na++ K+) activation and its significance for field properties

Abstract

AbstractA microsomal (Na++ K++ Mg2+)ATPase preparation from sugar beet roots was used. The activation by simultaneous addition of Na+ and K+ at different levels was examined in terms of steady state kinetics. The observed data can be summarized in the following way:1. The apparent affinity between the enzyme and the substrate MgATP depends on the ratio between Na+ and K+. At low Na+ concentration (below 5 mM), the apparent Km decreases with increasing concentrations of K+ (1–20 mM). At 5 mM Na+, the K+ level does not change the apparent Km, while at Na+ levels above 10 mM, the apparent Km between enzyme and substrate increases with increasing concentration of K+.2. When the MgATP concentration is kept constant, homotropic cooperativity (concerning one type of ligand) and heterotropic cooperativity (concerning different types of ligands) exist in the activation by Na+ and K+. The Na+ binding is cooperative with different Km values and Hill coefficients (n) in the presence of low and high concentration of K+. At low Na+ level (< 5 mM). a negative cooperativity exists for Na+ (nNa < 1) which is more pronounced in the presence of high [K+]. When the concentration of Na+ is raised the negative cooperativity disappears and turns into a positive one (nNa > 1).Only K+ binding in the presence of low [Na+] shows cooperativity with a Hill coefficient that reflects changes from negative to positive homotropic cooperativity with increasing concentrations of K+ (nK < 1 → nK > 1). In the presence of [Na+] > 10 mM, the changes in nk are insignificant.3. A model is proposed in which one or two different K sites and one or two Na sites control the catalytic activity, with multiple interactions between Na+, K+ and MgATP.4. In the presence of Na+ (< 10 mM), K+ is probably bound to two K sites, one of which translocates K+ through the membrane by an antiport Na+/K+ mechanism. This could be connected with an elevated K+ uptake in the presence of Na+ and could therefore explain some field properties of sugar beets.