Purification and characterization of glutamine synthetase from the commerical mushroom Agaricus bisporus |
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Authors: | J J P Baars H J M Op den Camp J W G Paalman V Mikeš C van der Drift L J L D Van Griensven G D Vogels |
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Institution: | (1) Department of Microbiology, Faculty of Science, University of Nijmegen, Toernooiveld 1, NL-6525 ED Nijmegen, The Netherlands;(2) Mushroom Experimental Station, Postbus 6042, NL-5960 AA Horst, The Netherlands;(3) Department of Biochemistry, Faculty of Science, Masaryk University, Kotlá ská 2, 61137 Brno, Czech Republic |
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Abstract: | Agaricus bisporus glutamine synthetase, a key enzyme in nitrogen metabolism, was purified to apparent homogeneity. The native enzyme appeared to be a GS-II type enzyme. It has a molecular weight of 325 kDa and consists of eight 46-kDa subunits. Its pI was found at 4.9. Optimal activity was found at 30°C. The enzyme had low thermostability. Stability declined rapidly at temperatures above 20°C. The enzyme exhibits a K
m for glutamate, ammonium, and ATP of 22mm, 0.16mm and 1.25mm respectively in the biosynthetic reaction, with optimal activity at pH 7. The enzyme is slightly inhibited by 10mm concentrations of l-alanine, l-histidine, l-tryptophan, anthranilic acid, and 5 -AMP and was strongly inhibited by methionine sulfoximine and phosphinothricine. For the transferase reaction K
i-values were 890 m and 240 m for methionine sulfoximine and phosphinothricine respectively. For the biosynthetic reaction K
i was 17 m for both methionine sulfoximine and phosphinothricine. |
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