Re-Evaluation of the Effects of Glucose-6-Phosphate and Malate on the Catalytic Properties of Phosphoenolpyruvate Carboxylase From Cynodon dactylon Under Physiological Assay Conditions

Abstract

Some catalytic and regulatory properties of the day- and night-form of phosphoenolpyruvate carboxylase (EC 4.1.1.31) from Cynodon dactylon (L.) Pers. leaves were re-evaluated in the presence of high concentrations of glycerol in the assay medium. This cosolute, known to be preferentially excluded from the hydration sphere of some proteins, apparently promotes aggregation, through an increase of the enzyme concentration in a fraction of the total volume. In the presence of this cosolute in the assay medium: (a) glucose-6-phosphate does not activate the enzyme, (b) inhibition of the V_max by malate is increased, particularly with the night-form of phosphoenolpyruvate carboxylase, whereas the day-form is more responsive at the K_m levels, (c) the light activation, detected as an increase in the apparent affinity of the substrate phosphoenolpyruvate is maintained under the new assay conditions.