Sawa Y, Ochiai H, Yoshida K, Tanizawa K, Tanaka H, Soda K
Laboratory of Biochemistry, College of Agriculture, Shimane University.
J Biochem. 1988 Dec;104(6):917-23. doi: 10.1093/oxfordjournals.jbchem.a122583.
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.
谷氨酰胺合成酶已从非固氮丝状蓝细菌拉氏席藻的细胞提取物中纯化至同质。通过十二烷基硫酸钠凝胶电泳测定该酶的亚基分子量约为59,000。拉氏席藻酶的电子显微镜照片显示出规则六边形的两层结构（每个分子12个亚基），这与通过X射线晶体学确定的鼠伤寒沙门氏菌谷氨酰胺合成酶的三维多肽主链结构非常相似（阿尔马西、扬森、哈姆林、熊和艾森伯格（1986年）《自然》323卷，304 - 309页）。拉氏席藻酶的N端氨基酸序列与固氮蓝细菌鱼腥藻7120的酶的N端氨基酸序列显示出非常高的相似性；在比较的23个残基中有18个残基相同。观察到针对纯化的拉氏席藻谷氨酰胺合成酶的抗体与除集胞藻酶之外的其他蓝细菌酶之间有强烈的免疫交叉反应。NH3、L - 谷氨酸和ATP的表观米氏常数分别测定为0.29、7.4和1.7 mM。二价金属离子如Mg2 +和Mn2 +在生物合成反应中激活该酶，而各种氨基酸和谷氨酸类似物强烈抑制该酶。
