Glutamine synthetase from a cyanobacterium, Phormidium lapideum: purification, characterization, and comparison with other cyanobacterial enzymes.
Glutamine synthetase has been purified to homogeneity from cell extracts of a non-N2-fixing filamentous cyanobacterium, Phormidium lapideum. The subunit molecular weight of the enzyme was determined as about 59,000 by sodium dodecyl sulfate gel electrophoresis. Electron micrographs of the Phormidium enzyme revealed a two-layered structure of regular hexagons (12 subunits per molecule), which markedly resembles the three-dimensional polypeptide backbone structure of the Salmonella typhimurium glutamine synthetase established by X-ray crystallography (Almassy, Janson, Hamlin, Xuong, & Eisenberg (1986) Nature 323, 304-309). The N-terminal amino acid sequence of the Phormidium enzyme shows very high similarity with that of the enzyme from an N2-fixing cyanobacterium, Anabaena 7120; 18 residues are common in 23 residues compared. Strong immunocross-reactions between the antibody against the purified Phormidium glutamine synthetase and other cyanobacterial enzymes except the Anacystis enzyme were observed. The apparent Michaelis constants for NH3, L-glutamate, and ATP were determined to be 0.29, 7.4, and 1.7 mM, respectively. Divalent metal ions such as Mg2+ and Mn2+ activated the enzyme in the biosynthetic reaction, whereas various amino acids and glutamate analogs strongly inhibited the enzyme.
Sawa
Y
Y
Laboratory of Biochemistry, College of Agriculture, Shimane University.
Ochiai
H
H
Yoshida
K
K
Tanizawa
K
K
Tanaka
H
H
Soda
K
K
D000595:Amino Acid Sequence; D002413:Cations, Divalent; D000458:Cyanobacteria / Q000201:enzymology*; D005974:Glutamate-Ammonia Ligase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007120:Immunochemistry; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D015394:Molecular Structure; D008970:Molecular Weight; D011487:Protein Conformation
D002413:Cations, Divalent; D005974:Glutamate-Ammonia Ligase