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| TITLE | Purification and characterization of glutamine synthetase from the unicellular cyanobacterium Synechococcus RF-1 |
| AUTHOR | Hso-Freng Yuan* Institute of Botany, Academia Sinica, Taipei, Taiwan 11529, Republic of China Chiung-Min Wang Institute of Botany, Academia Sinica, Taipei, Taiwan 11529, Republic of China Han-Wei Kung Institute of Botany, Academia Sinica, Taipei, Taiwan 11529, Republic of China |
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| ABSTRACT | Glutamine synthetase (GS; EC 6.3.1.2) from the Synechococcus RF-1 was purified to homogeneity by ion exchange, molecular sieving, and hydroxyapatite chromatographies. The native enzyme has a molecular mass of about 456 kDa, and the molecular mass of its subunit was about 56 kDa. Electron micrographs of the enzyme revealed two parallel protein layers in cubic symmetry with quaternary structure. The actual data indicated that the enzyme could consist of eight identical subunits. The enzyme had an apparent Km value for L-glutamate of 2.33 mM, but it exhibited positive cooperativity for ATP (nH = 1.5 and S0.5 = 0.94 mM) and NH4Cl (nH = 2, and S0.5 = 1.33 mM) in the biosynthetic assay. The enzyme had apparent Km values for L-glutamine and hydroxylamine of 8.70 mM and 7.04 mM, respectively, in the transferase assay. This enzyme was quite stable in Tris-HCl buffer (pH 7.5) containing EDTA, MgCl2 and 2-mercaptoethanol. The pH optima for both the biosynthetic and transferase activities of the enzyme were 8.1 and 8.4, respectively. The enzyme required a divalent metal ion as an activator. Mg2+ was the most effective metal ion for biosynthetic activity, followed by Co2+. Mn2+ was the most effective metal ion for transferase activity. Ca2+ and Mn2+ strongly inhibited Mg2+-supported biosynthetic activity, but Co2+ stimulated it. |
| KEYWORD | Cyanobacteria; Glutamine synthetase; Synechococcus RF-1; |
| ARTICLE INFO | Botanical Bulletin of Academia Sinica, Volume 42 Number 1 January 2001, page 23-33, 11 pages |
| PUBLISHER | Institute of Plant and Microbial Biology, Academia Sinica, Taipei, Taiwan, Republic of China |
