Study of the effect of lanthanide ions on the kinetics of glutamate dehydrogenase by a chronoamperometric method

Abstract

A chronoamperometric method, based on the oxidation of reduced nicotinamide adenine dinucleotide (NADH) on a glassy carbon electrode, for assaying glutamate dehydrogenase activity is described. Once the reaction was initiated by addition of enzyme into a well stirred assay mixture containing NADH, the concentration of NADH and the steady-state limiting current of NADH oxidation on the electrode decreased linearly in a short time range from the start of the reaction. The rate of the enzyme-catalysed reaction can be calculated from the decrease in the current. This method is reliable for the accurate determination of enzyme activity in purified extracts from liver and crude extracts from cucumber root. The major advantage of this method is the real-time and continuous measurement of the change in coenzyme concentration, so the real initial reaction rate, ν₀, can be determined. This method has been used to detect the effect of La³⁺, Eu³⁺, Yb³⁺ and Zn²⁺ ions on the enzyme activity. The results suggest that lanthanide ions can increase the activity of the enzyme in glutamate synthesis, and Zn²⁺ acts as a potent inhibitor of the enzyme.