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Ammonium assimilation in bryophytes. l ‐glutamine synthetase from Sphagnum fallax
Author(s) -
Kahl Stefan,
Gerendás Jóska,
Heeschen Volker,
Ratcliffe R. George,
Rudolph Hansjörg
Publication year - 1997
Publication title -
physiologia plantarum
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.351
H-Index - 146
eISSN - 1399-3054
pISSN - 0031-9317
DOI - 10.1111/j.1399-3054.1997.tb01823.x
Subject(s) - glutamine synthetase , ammonium , biochemistry , nitrogen assimilation , azaserine , molecular mass , enzyme , amino acid , biology , size exclusion chromatography , glutamine , chemistry , organic chemistry
Cytosolic and plastidic l ‐glutamine synthetase (EC 6.3.1.2) isoenzymes from Sphagnum fallax Klinggr. (Klinggr. clone 1) were separated by size‐exclusion and ion exchange chromatography. The cytosolic enzyme (GS 1 ) was purified to apparent electrophoretic homogeneity. The native enzyme had a molecular mass of 390 ± 20 kDa as estimated by gel filtration and was apparently composed of 8 subunits with molecular masses of 48 kDa. GS 1 activity could be measured from pH 6.8 to 8.6 in 50 m M imidazole buffer, with a broad optimum between pH 7.2 and 8.0. The K m values were 2.5 m M , 0.5 m M and 0.5 m M for l ‐glutamate, ammonium and ATP, respectively. The enzyme was inhibited by more than 10 m M ammonium or glutamate. The incorporation of 15 NH 4 + into amino acids was observed in vivo using 15 NMR. Label from ammonium was first detected in the amide N of glutamine, and only subsequently in the amino N of glutamate. Moreover, no assimilation was detected in the presence of the specific GS inhibitor methionine sulfoximine. These observations are consistent with a dominant role for GS in the assimilation of ammonium in Sphagnum .