Kinetic Studies on Na+/K+‐ATPase by Using Thermokinetic Method

Abstract

AbstractNa+/K−‐ATPasc (EC 3.6.1.3) is an important membrane‐bound enzyme. By using microcalorimetry, the thermokinetic method was developed to kinetic studies on Na+/K+‐ATPase for the first time. Compared with other ones, the method provided accurate measurements of not only thermodynamic data but also the kinetic data. At 310.15 K and pH=7.4, the molar reaction enthalpy ΔrHm, was measured as (‐40.408k 1.9) kJ mol−1. The Michaelis constant Km was determined to be (0.479 0.020) 10−‐3 mol L−1 and consistent with literature figure which is about 0.5 10−‐3 moi L−1. The maximum velocity Vmax obtained was (0.6810.026) μmol Pi min−1 mg protein−1. All of the data have good repeatability and self‐consistency. The reliability of thermokinetic method was verified by the experimental results and further confirmed by colorimetric studies. Moreover. the effect of enzyme pre‐dilution on its activities was also investigated.