Activation of Kidney Mitochondrial Glutaminase by Inorganic Phosphate and Organic Acids

Abstract

THE deamidation of glutamine to glutamic acid and ammonia by animal tissues is accelerated by phosphate and certain other divalent inorganic anions^1,2. Of the organs studied, the kidney has the highest concentration of the enzyme which catalyses this reaction³. A tentative mechanism for the action of renal glutaminase has been proposed⁴ on the basis of its activation by phosphate. The active centre of the enzyme is thought to have at least two cationic sites and possibly one sulphydryl site⁵. This communication presents the results of experiments showing the acceleration of glutamine hydrolysis in kidney tissue by a group of carboxylic acids most of which are intermediates of the Krebs tricarboxylic acid cycle. Kinetic studies reveal similarities between the activation of glutaminase by these acids and that by inorganic phosphate. Kidney cortices from rats were homogenized in ice cold 0.154 molar potassium chloride at 4° C. Mitochondria were prepared by spinning the homogenate at 800g for 15 min and then re-spinning the supernatant at 12,000g for 20 min. The mitochondrial pellet was washed twice. The washed mitochondria were then incubated with glutamine in tris buffer at 37° C. When salts of the organic acids and phosphate were added, their concentration in the incubation medium, unless otherwise stated, was always 0.3 molar. The pH was 8.0 unless varied as stated.