Acetylglutamate kinase: a feedback-sensitive enzyme of arginine biosynthesis in Neurospora.

Jan J. Cybis* and Rowland H. Davis Department of Botany, University of Michigan, Ann Arbor, Michigan 48104 Received August 6,1974 summary: A radioactive assay was developed for the arginine-synthetic enzyme, acetylglutamate kinase (EC 2.7.2.8). Activity of the enzyme was demonstrated in crude extracts of Neurospora mycelium. Precipitation with ammonium sulfate, resulting in separation of the enzyme from an inhibitor, was initially required to detect activity. Most preparations are only partially sensitive to arginine, with maximal inhibition achieved at an effector concentration of 0.5 mM. The enzyme is activated about 10% by 1 mM lysine or citrulline, while 1 mM ornithine stimulates activity by 75%. The compartmsntation of the arginine pool (1,Z) and the mitochondrial location of some arginine biosynthetic enzymes (3) of Neurospora makes the question of feedback inhibition in the pathway especially interesting. Feedback inhibition of omithine synthesis by arginine was postulated on physiological grounds earlier (4). Omithine synthesis begins with formation of acetylglutamate from glutamate by an acetyl-CoA-dependent enzyme. However, in Neurospora, acetylglutamate is also regenerated in a transfer of the acetyl group of Nd-acetylornithine to glutamate, which completes a cycle in the reaction liberating ornithine. It was expected, therefore, that a key feedback-sensitive enzyme would be acetylglutamate kinase, which transforms acetylglutamate to N-acetyl-J-glutamyl phosphate. Earlier attempts to demonstrate the enzyme in Neurospora (5) were unsuccessful. A dependable assay for the kinase was sought to determine its sensitivity to arginine and to facilitate studies of repression and localization.