Nitrogen and ammonia assimilation in the cyanobacteria: purification of glutamine synthetase from Anabaena sp. strain CA.
Glutamine synthetase was purified from the cyanobacterium Anabaena sp. strain CA, a newly isolated marine organism. This organism grows rapidly under nitrogen-fixing conditions and therefore is ideally suited for studies concerning cyanobacterial nitrogen metabolism. Studies were conducted to optimize the production of glutamine synthetase by Anabaena CA. The highest specific activities were obtained from cells grown in the presence of atmospheric N(2) or KNO(3) (13 mM); when NH(4)Cl was used as the nitrogen source, the specific activity was reduced by approximately 40%. Furthermore, through the use of a whole-cell gamma-glutamylhydroxamate transferase assay, it was found that the maximum number of enzyme units is obtained in the late logarithmic stage of growth. Glutamine synthetase purification requires only three steps and results in a preparation that is electrophoretically homogeneous. The transferase specific activity (units per milligram of protein) of the purified enzyme is 78, whereas the biosynthetic specific activity is 2.2. The molecular weight of the native protein was found to be approximately 590,000, and the subunit molecular weight was determined to be about 50,000. Thus, this cyanobacterial enzyme closely resembles the enzyme obtained from other procaryotic sources, at least with regard to size. The purification of glutamine synthetase from Anabaena CA should stimulate a more detailed study of this enzyme and its role in cyanobacterial nitrogen metabolism.
D000641:Ammonia / Q000378:metabolism*; D000458:Cyanobacteria / Q000201:enzymology / Q000254:growth & development / Q000378:metabolism*; D005974:Glutamate-Ammonia Ligase / Q000096:biosynthesis / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D009584:Nitrogen / Q000378:metabolism*; D012623:Seawater; D014871:Water Microbiology
D000641:Ammonia; D005974:Glutamate-Ammonia Ligase; D009584:Nitrogen