Kinetic and thermodynamic study of the reaction catalyzed by glucose-6-phosphate dehydrogenase with nicotinamide adenine dinucleotide

The enzyme glucose-6-phosphate dehydrogenase (G6PD, EC 1.1.1.49) from Leuconostoc mesenteroides has a dual coenzyme specificity with oxidized nicotinamide adenine dinucleotide (NADox) and oxidized nicotinamide adenine dinucleotide phosphate as electron acceptors. The G6PD coenzyme selection is determined by the metabolic cellular prevailing conditions. In this study a kinetic and thermodynamic analysis is presented for the reaction catalyzed by G6PD from L. mesenteroides with NADox as coenzyme in phosphate buffer. For this work, an in situ spectrophotometric technique was employed based on the detection of one product of the reaction. Substrate and coenzyme concentrations as well as temperature and pH effects were evaluated. The apparent equilibrium constant, the Michaelis constant, and the turnover number were determined as a function of each experimental condition. The standard transformed Gibbs energy of reaction was determined from equilibrium constants at different initial conditions. For the product 6-phospho-d-glucono-1,5-lactone, a value of the standard Gibbs energy of formation is proposed, ΔfG° = −1784 ± 5 kJ mol−1.

Research highlights
► The reaction catalyzed by one enzyme of the pentose phosphate pathway was studied.
► A spectrophotometric method is proposed for kinetic and thermodynamic analysis.
► The pH and the temperature influences are reported on physical chemical properties.
► Relative concentrations of substrates are also important in the catalytic process.