Properties of rat kidney glutaminase enzymes and their role in renal ammoniagenesis

Abstract

Rat kidney contains two distinct glutaminase activities; the mitochondrial phosphate-dependent glutaminase and a second glutaminase activity associated with the brush border membrane which is maleate-activated and phosphate-independent. It has recently been shown that the phosphate-independent glutaminase is a partial reaction of gamma-glutamyl transpeptidase and that maleate activates this enzyme by blocking transpeptidation. The gamma-glutamyl transpeptidase in other rat tissues is also affected by maleate. This enzyme has at least a 100-fold greater affinity for glutathione or for glutathione derivatives than for glutamine, suggesting that under physiological conditions glutathione is the preferred substrate. With either type of substrate, maleate affects the Vmax of the reaction but not the Km. These findings suggest that this enzyme probably contributes very little to renal ammoniagenesis. In contrast, the phosphate-dependent glutaminase, whose activity increases 20 to 30-fold in the proximal convoluted tubule cells in response to metabolic acidosis, probably contributes significantly to renal ammoniagenesis. We have purified the rat kidney phosphate-dependent glutaminase and compared the phosphate activation and the phosphate-induced dimerization of the Tris form of this enzyme. There is an excellent correlation between increased activity and extent of dimerization as phosphate concentration is increased. The molecular weights of the Tris form are 1600000 and 316000 in the absence and presence of -1 M NaPO4, respectively. At saturating concentration of phosphate, increasing concentrations of chloride ion similarly reverse both activation and dimerization. These observations suggest that only the dimer form of the Tris enzyme is active.