Ammonia Assimilation by Rhizobium Cultures and Bacteroids
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Abstract:
The enzymes involved in the assimilation of ammonia by free-living cultures of Rhizobium spp. are glutamine synthetase (EC. 6.o.I.2), glutamate synthase (L-glutamine:2-oxoglutarate amino transferase) and glutamate dehydrogenase (ED I.4.I.4). Under conditions of ammonia or nitrate limitation in a chemostat the assimilation of ammonia by cultures of R. leguminosarum, R. trifolii and R. japonicum proceeded via glutamine synthetase and glutamate synthase. Under glucose limitation and with an excess of inorganic nitrogen, ammonia was assimilated via glutamate dehydrogenase, neither glutamine synthetase nor glutamate synthase activities being detected in extracts. The coenzyme specificity of glutamate synthase varied according to species, being linked to NADP for the fast-growing R. leguminosarum, R. melitoti, R. phaseoli and R. trifolii but to NAD for the slow-growing R. japonicum and R. lupini. Glutamine synthetase, glutamate synthase and glutamate dehydrogenase activities were assayed in sonicated bacteroid preparations and in the nodule supernatants of Glycine max, Vicia faba, Pisum sativum, Lupinus luteus, Medicago sativa, Phaseolus coccineus and P. vulgaris nodules. All bacteroid preparations, except those from M. sativa and P. coccineus, contained glutamate synthase but substantial activities were found only in Glycine max and Lupinus luteus. The glutamine synthetase activities of bacteroids were low, although high activities were found in all the nodule supernatants. Glutamate dehydrogenase activity was present in all bacteroid samples examined. There was no evidence for the operation of the glutamine synthetase/glutamate synthase system in ammonia assimilation in root nodules, suggesting that ammonia produced by nitrogen fixation in the bacteroid is assimilated by enzymes of the plant system.Keywords:
Glutamate synthase
Nitrogen Assimilation
Rhizobium leguminosarum
The enzymes involved in the assimilation of ammonia by free-living cultures of Rhizobium spp. are glutamine synthetase (EC. 6.o.I.2), glutamate synthase (L-glutamine:2-oxoglutarate amino transferase) and glutamate dehydrogenase (ED I.4.I.4). Under conditions of ammonia or nitrate limitation in a chemostat the assimilation of ammonia by cultures of R. leguminosarum, R. trifolii and R. japonicum proceeded via glutamine synthetase and glutamate synthase. Under glucose limitation and with an excess of inorganic nitrogen, ammonia was assimilated via glutamate dehydrogenase, neither glutamine synthetase nor glutamate synthase activities being detected in extracts. The coenzyme specificity of glutamate synthase varied according to species, being linked to NADP for the fast-growing R. leguminosarum, R. melitoti, R. phaseoli and R. trifolii but to NAD for the slow-growing R. japonicum and R. lupini. Glutamine synthetase, glutamate synthase and glutamate dehydrogenase activities were assayed in sonicated bacteroid preparations and in the nodule supernatants of Glycine max, Vicia faba, Pisum sativum, Lupinus luteus, Medicago sativa, Phaseolus coccineus and P. vulgaris nodules. All bacteroid preparations, except those from M. sativa and P. coccineus, contained glutamate synthase but substantial activities were found only in Glycine max and Lupinus luteus. The glutamine synthetase activities of bacteroids were low, although high activities were found in all the nodule supernatants. Glutamate dehydrogenase activity was present in all bacteroid samples examined. There was no evidence for the operation of the glutamine synthetase/glutamate synthase system in ammonia assimilation in root nodules, suggesting that ammonia produced by nitrogen fixation in the bacteroid is assimilated by enzymes of the plant system.
Glutamate synthase
Nitrogen Assimilation
Rhizobium leguminosarum
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summary In order to investigate the relative contribution of glutamine synthetase and NADP‐glutamate dehydrogenase to the assimilation of ammonium (NH 4 + ) by spruce ectomycorrhizas, changes in free amino acid content and kinetics of 15 N incorporation into free amino acids were measured together with the effect of specific enzyme inhibitors. Exposure of detached ectomycorrhizas to ( 15 NH 4 ) 2 SO 4 showed that the greatest flow of 15 N enters into the amido group of glutamine. Label was also detected in glutamic acid, alanine and γ‐aminobutyric acid. Large amounts of alanine and glutamate accumulated in response to the addition of methionine sulfoximine (MSX) together with a decrease in 15 N incorporation into both amido‐ and ammo‐nitrogen of glutamine. These results are consistent with a major role of glutamate dehydrogenase and glutamine synthetase in nitrogen assimilation in the symbiosis and do not suggest any significant role for glutamate synthase in the synthesis of glutamate. A large accumulation of unlabelled asparagine in response to MSX and albizziine inhibition suggests the occurrence of an unlabelled NH 4 + pool in the host plant. The transfer of nitrogen compounds between the fungal cells and the host tissues is discussed.
Glutamate synthase
Nitrogen Assimilation
Asparagine synthetase
Alanine
Glutamic acid
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SUMMARY The N assimilation pathways in leaves of Datura stramonium L. have been investigated at low and high levels of xylem stream nitrate feeding by an 15 N infiltration technique in the presence and absence of the glutamine synthetase inhibitor, methionine sulphoximine (MSO). In the absence of MSO, glutamate showed the highest 15 N enrichment of any free amino compound at both feeding levels. At the high N feeding level, glutamine showed the highest accumulation of 15 N at all stages of the time course, with secondary accumulation in glutamate. At the low N feeding level the reverse was true. These findings indicate the operation of both the glutamine synthetase‐GOGAT and glutamate dehydrogenase pathways in Datura leaves. In the presence of MSO intracellular pools of all the amino compounds, with the exception of glutamine and threonine, became rapidly drained. There was an almost complete restriction of the supply of newly reduced 15 N to amino acid assimilation and it accumulated as ammonia. The large glutamine pool remaining contained no 15 N enrichment, and was probably an extrachloroplastic storage pool not immediately available for amino acid synthesis. These experimental results indicate the almost exclusive role of the glutamine synthetase‐GOGAT pathway in the assimilation of newly reduced nitrate into amino acid metabolism in Datura leaves.
Glutamate synthase
Datura stramonium
Nitrogen Assimilation
Azaserine
Glufosinate
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Glutamate synthase
Nitrogen Assimilation
Metabolic pathway
Assimilation (phonology)
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Regulation of the enzymes involved in glutamine and glutamate biosynthesis in Bacillus spp and their roles in ammonia assimilation is the focus of this chapter. For most bacteria, assimilation of ammonia is accomplished through the synthesis of glutamine and glutamate. Alanine dehydrogenase and asparagine synthetase have also been implicated in assimilation, and their roles are discussed. The Bacillus spp can be separated into three groups based on the pathway used for assimilation. One group, represented by B. subtilis, employs glutamine synthetase (GS) and GOGAT for assimilation. The second, which contains most members of the genus, utilizes all three enzymes, depending on nutritional environment. The third group uses only GDH for assimilation, a characteristic of some N2-fixing Bacillus spp. Assimilation of ammonia in derivatives of B. subtilis 168 and SMY is solely accomplished through the coupled action of GS and GOGAT. A catabolic role for GDH is consistent with the notion that GOGAT is solely responsible for glutamate biosynthesis in B. subtilis. The production of glutamate from glutamine is accomplished by the enzyme L-glutamine aminohydrolase, also known as glutaminase.
Glutamate synthase
Glutaminase
Assimilation (phonology)
Nitrogen Assimilation
Asparagine synthetase
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Asparagine synthetase
Nitrogen Assimilation
Glutamate synthase
Assimilation (phonology)
Azaserine
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Assimilation of nitrate and ammonium are vital procedures for plant development and growth. From these primary paths of inorganic nitrogen assimilation, this metabolism integrates diverse paths for biosynthesis of macromolecules, such as amino acids and nucleotides, and the central intermediate metabolism, like carbon metabolism and photorespiration. This paper reports research performed in the CitEST (Citrus Expressed Sequence Tag) database for the main genes involved in nitrogen metabolism and those previously described in other organisms. The results show that a complete cluster of genes involved in the assimilation of nitrogen and the metabolisms of glutamine, glutamate, aspartate and asparagine can be found in the CitEST data. The main enzymes found were nitrate reductase (NR), nitrite reductase (NiR), glutamine synthetase (GS), glutamate synthetase (GOGAT), glutamate dehydrogenase (GDH), aspartate aminotransferase (AspAT) and asparagine synthetase (AS). The different enzymes involved in this metabolism have been shown to be highly conserved among the Citrus and Poncirus species. This work serves as a guide for future functional analysis of these enzymes in citrus.
Glutamate synthase
Nitrogen Assimilation
Asparagine synthetase
Photorespiration
Nitrogen Cycle
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The regulation of glutamate dehydrogenase (EC 1.4.1.4), glutamine synthetase (EC 6.3.1.2), and glutamate synthase (EC 2.6.1.53) was examined for cultures of Salmonella typhimurium grown with various nitrogen and amino acid sources. In contrast to the regulatory pattern observed in Klebsiella aerogenes, the glutamate dehydrogenase levels of S. typhimurium do not decrease when glutamine synthetase is derepressed during growth with limiting ammonia. Thus, it appears that the S. typhimurium glutamine synthetase does not regulate the synthesis of glutamate dehydrogenase as reported for K. aerogenes. The glutamate dehydrogenase activity does increase, however, during growth of a glutamate auxotroph with glutamate as a limiting amino acid source. The regulation of glutamate synthase levels is complex with the enzyme activity decreasing during growth with glutamate as a nitrogen source, and during growth of auxotrophs with either glutamine or glutamate as limiting amino acids.
Glutamate synthase
Glutamic acid
Glutaminase
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Maize growth and ammonium assimilation enzyme activity in response to nitrogen forms and pH control1
Abstract Plant growth, glutamine synthetase and glutamine dehydrogenase activities of two maize genotypes were compared in the presence of NH4 + and NO3 ‐ forms of N in sand culture. Ammonium reduced growth of the P3732 genotype 64% and the B73 x Mol7 hybrid 59% as compared to NO3 ‐. Both glutamine synthetase and glutamate dehydrogenase activities in roots tended to be higher with NH4 as compared to NO3. As the pH in the medium was increased by adding CaCO3, glutamine synthetase and glutamate dehydrogenase activities in roots of both genotypes were reduced; however, glutamine synthetase activity in leaves of NH^‐treated plants increased at the higher pH of the growing medium.
Glutamate synthase
Nitrogen Assimilation
Assimilation (phonology)
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summary In order to determine significance of the glutamate dehydrogenase pathway and a glutamine synthetase/glutamate synthase cycle in NH 4 + assimilation, we followed a number of different metabolikc parameters (nutrient uptake, free amino acid pools, NH 4 + ‐induced glutamine accumulation) and 15 N incorporation into amino acids of rapidly growing Cennoccum geophilum. Arginine was a major free amino acid in C. geophilum during its entire growth period. C. geophilum synthesized and accumulated very large amounts of glutamine at the beginning of the rapid phase of growth in low nitrogen medium, during the whole growth period in high nitrogen medium, and immediately after addition of NH 4 + . Therefore, the accumulation of a large amount of glutamine tool; place when the external ammonium concentration was high. The current data identify four pathways of N metabolism in rapidly growing C. geophilum : (1) glutamine synthesis, invoking transfer of N to both amino and amino moieties; (2) glutamate formation; (3) transamination with pyruvate to yield alanine; (4) transamination with oxaloacetate to yield asparate. The higher accumulation of glutamate and related amino acids (alanine and aspartate) in the presence of the glutamine synthetase inhibitor methionine sulphoximine indicates that glutamate, the precursor of glutamine, was formed by a pathway insensitive to methionine sulphoximine, the glutamate dehydrogenase pathway. Up to 40% of the assimilated 15 N terminated in the amido‐N of glutamine. These data are consistent with a pivotal role for glutamine synthetase activity and indicates that the primary assimilation of NH 4 + in rapidly growing C. geophilum is brought about by concurrent activity of the GDH and GS pathways. The pathway of primary assimilation of NH 4 + by C. geophilunt in the rapid phase of growth therefore differs from those operating in the stationary phase of growth where N flax through GDH is higher than the flux through the GS pathway.
Glutamate synthase
Transamination
Nitrogen Assimilation
Amino acid synthesis
Alanine
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