Influence of Ammonium and Nitrate Nutrition on the Pyridine and Adenine Nucleotides of Soybean and Sunflower

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Energy charge I(ATP) + 1/2 (ADP)J/[(ATP) + (ADP) + (AMP)]and glutamine synthetase activity (transferase reaction) of roots increase in a near congruent manner when decotyledonized sunflower plants (Helianthus annuus L. var. Mammoth Russian) are grown in nitrate for 9 days. Replacement of nitrate with ammonium for the final 2 days leads to a higher energy charge and increased enzyme activity. Similar (20,23,28). An enzyme utilizing the energy of ATP in bond formation is glutamine synthetase (EC 6.3.1.2.). Responsible as it is for the formation of glutamine from glutamic acid, ammonia, and ATP, the enzyme is also known to function in vitro as a glutamyl transferase (19). Evidence has previously been presented indicative of a correlation between the energy charge of soybean and sunflower roots and the glutamyl transferase activity of these tissues (28). In a further effort to investigate the relationship between energy charge and the specific activity of a biosynthetic enzyme, the present study has examined glutamine synthetase activity (transferase reaction) and adenosine phosphate content in sunflower root tissue of plants supplied with nitrate after removal of cotyledons, of plants transferred from a nitrate culture solution to an ammonium solution, of plants subjected to a zero nitrogen medium, and of plants treated to an extended period of darkness.In microorganisms (31) and in rice roots (12) transferase activities of the enzyme have been demonstrated as sensitive to inhibition by various amino acids generally viewed as end products of glutamine metabolism. Accordingly, the influence of amino acids in the regulation of glutamine synthetase has been examined in this study by a determination of the concentration in the root tissues of a possible end product, alanine, together with the levels of glutamic acid and glutamine. MATERIALS AND METHODSPlant Culture. Sunflower seeds (Helianthus annuus L. var. Mammoth Russian) were germinated in filter paper-lined beakers for a period of 5 days. In experiments in which only nitrate was to be utilized as a nitrogen source, pairs of seedlings were transferred to 600-ml beakers containing culture solution and were supported with roots immersed in solution by doublegrooved plastic beaker covers. Culture solution for these plants was changed daily. For the experiment in which ammonium was provided after a period of nitrate culture, 5-day-old seedlings were transferred to the continuous flow apparatus previously described (29), and culture solution was passed over the roots for the remainder of the 20-day growth period. Under these conditions of growth, culture solutions experienced no more than a decrease of 0.1 pH unit during a 24-hr period.Following the transfer to either 600-ml beakers or to the continuous flow apparatus, all plants were treated with culture solution lacking in nitrogen (25) for a period of 7 days. The cotyledons were then removed from the 12-day plants, and culture solution containing 10 mm KNO3 was provided. All plants were grown in a contro...

Section: Abstract Energy Charge I(atp) + 1/2 (Adp)j/[(atp) + (Adp) + supporting

confidence: 48%

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confidence: 99%

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confidence: 99%

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Glutamine Synthetase Regulation by Energy Charge in Sunflower Roots

Weissman

1976

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“…Additional determining factors may be the accumulation of amino acids in the metabolic pool (23) and the availability in root tissue of reducing power in the form of NADH and NADPH. The question is further examined in an accompanying study (31).…”

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confidence: 99%

Influence of Ammonium and Nitrate Nutrition on Enzymatic Activity in Soybean and Sunflower

Weissman

1972

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Under conditions of controlled pH, nitrate and ammonium are equally effective in supporting the growth of young soybean (Glycine max var. Bansei) and sunflower (Helianthus annuus L. var., Mammoth Russian) plans. Soybean contains an active nitrate reductase in roots and leaves, but the low specific activity of this enzyme in sunflower leaves indicates a dependency upon the roots for nitrate reduction. Suppression of nitrate reductase activity in sunflower leaves may be due to high concentrations of ammonia received from the roots. Nitrate reductase activity in leaves of nitrate-supplied soybean and sunflower follows closely the distribution of nitrate reductase. For the roots of both species, glutamic acid dehydrogenase activity was greater with ammonium than with nitrate.The glutamic acid dehydrogenase of ammonium roots is wholly NADH-dependent, whereas that of nitrate roots is active with NADH and NADPH. In leaves, an NADPH-dependent glutamic acid dehydrogenase appears to be responsible for the assimilation of translocated ammonia and ammonia formed by nitrate reduction.In soybean roots ammonia is actively incorporated into amides, much of which remains in the roots. Sunflower roots are less active in amide formation but transfer much of it, together with ammonia, into the shoots. Glutamine synthetase activity in leaves is 20-to 40-fold lower than in roots.Glucose-6-phosphate dehydrogenase activity appears to be correlated with the activity of the nitrate reducing system in roots, but not in leaves.Although it is well established that absorbed nitrate is converted to ammonium before assimilation into the organic form, it has long been recognized that ammonium and nitrate as sources of external nitrogen differ in their effects on the growth and chemical composition of plants (19). Recent In initial experiments sunflower seeds (Helianthus annuus L. var. Mammoth Russian) and soybean (Glycine max var. Bansei) were germinated in filter-paper lined beakers and were transferred as 5-day-old seedlings to sand-filled paraffined cups as previously described (30). Culture solution lacking in nitrogen (29) was provided for the first 10 days of growth. The cotyledons were then removed, and culture solution containing either 10 mm NH4C1 or 10 mm KNO3 was added twice a day for the final 9 days of growth before harvest. Examination of the drippings passing through the glass wool covered perforations in the bottom of the cups on the final day of growth revealed that ammonium solutions, originally at pH 6.3, fell to pH 5.3 while nitrate solutions fell to pH 6.1. In order to minimize the effect of pH differences on the biochemical characteristics of ammonium-supplied and nitratesupplied plants, an Homogenates for the analysis of GDH activity were prepared by the method of Ritenour et al. (20). After the slurry was filtered through cheesecloth, the filtrate was centrifuged at 200g, the sediment was discarded, and the supernatant was centrifuged at 20,000g. Following the procedure described by Leech and Kirk (16), the p...

“…Reduction of N03 to NH4, consumes reducing equivalents and the ratio NADPH/NADP+ in leaves of NH4+-fed plants has been shown to be higher than in plants supplied with N03- (28). NADPH, but not NADP+, apparently stimulates RuBP carboxylase activity (5) and it may be that the turnover of carbon through the PCR cycle is partly regulated by the N source through the relative levels of oxidized and reduced nucleotides.…”

Section: Discussionmentioning

confidence: 99%

Effects of Different Inorganic Nitrogen Sources on Photosynthetic Carbon Metabolism in Primary Leaves of Non-nodulated Phaseolus vulgaris L.

Marques

Oberholzer²,

Erismann³

1983

Young bean plants (Phaseolus vulgaris L. var Saxa) were fed with three different types of inorganic nitrogen, after being grown on nitrogen-free nutrient solution for 8 days. The pattern of "CO2 fixation was investigated in photosynthesizing primary leaf discs of 11-day-old plants (3 days with nitrogen source) and in a pulse-chase experiment in 13-day-old plants (5 days with nitrogen source).Ammonium caused, in contrast to nitrate nutrition, a higher level of 14C incorporation into sugar phosphates but a lower incorporation of label into malate, glycolate, glycerate, aspartate, and alanine. The labeling kinetics of glycine and serine were little changed by the nitrogen source. Ammonium feeding also produced an increase in the ratio of extractable activities of ribulose-1,5-bisphosphate carboxylase to phosphoenolpyruvate carboxylase and an increase in dark respiration and the CO2 compensation concentration. Net photosynthesis was higher in plants assimilating nitrate.The results point to stimulated turnover of the photosynthetic carbon reduction cycle metabolites, reduced phosphoenolpyruvate carboxylation, and altered turnover rates within the photosynthetic carbon oxidation cycle in ammonium-fed plants. Mechanisms of the regulation of primary carbon metabolism are proposed and discussed.The effect of NH4' on photosynthetic carbon assimilation has been repeatedly reported (3, 8, 12, 13, 16, 17, 19-23, 27, 29) a decrease in the concentration of malate and an inhibition of malic enzyme activity (2). However, the supply of N03-to cells of Spinacia oleracea had no apparent effect on the distribution of photosynthetic products compared with N-free nutrition, whereas in NH4t-fed cells, fixation of CO2 into carbohydrates decreased but increased in carboxylic and amino acids (16).The effect of inorganic N sources on the composition of N compounds in the bleeding sap and leaves of the bushbean Phaseolus vulgaris has previously been reported (6, 27). Coupled with the above observations, this has led us to investigate the effect of different N sources on various carboxylating enzymes and on the pattern of "4CO2 fixation in primary leaves of nonnodulated bean plants grown in liquid culture. Our results show that different N sources, fed through the roots of Phaseolus vulgaris, cause changes in photosynthetic carbon metabolism. MATERIALS AND METHODSPlant Material. Seeds of Plaseolus vulgaris L. var Saxa (strain Vatter) were soaked overnight in tap water, rinsed, and placed on wet tissue paper in darkness for 3 d at 24 ± 2°C. Seedlings were transferred to coarse quartz sand, watered with N-free nutrient (27), and grown under an illumination of 7000 to 9000 lux. After a further 4 d (day 8), the plants were set into plastic tanks filled with 14 L nutrient solution containing a total N concentration of 3.5 mm either as NH4', or NH4NO3 (27). The pH of the nutrient solution was adjusted with either KOH or H3PO4. All subsequent investigations were carried out on primary leaves. "4C02/'2C02 Pulse-Chase ExpeAments. After...