Purification, Properties and Regulation of Glutamine Synthetase from Nitrobacter agilis

Summary: Glutamine synthetase (EC 6.3.1.2) was purified about 430-fold from the nitrifying bacterium Nitrobacter agilis by affinity chromatography on Blue Sepharose CL-6B and gel filtration on Sepharose 4B. The enzyme (apparent mol. wt 700000), which consisted of 12 subunits, each of mol. wt 58000, required a divalent cation for both biosynthetic and transferase activities. Regulation of glutamine synthetase both by a feedback inhibition involving amino acids and by an adenylylation/deadenylylation mechanism was studied. The enzyme was highly adenylylated (75-90%) in cell free extracts from cells grown on nitrite as the sole source of nitrogen. The adenylylated form of the enzyme could be deadenylylated by treatment with snake venom phos-phodiesterase. An isoactivity pH of 7·4 was recorded for glutamine synthetase.