Alfalfa Root Nodule Carbon Dioxide Fixation

Miller, Stephen R.; Boylan, Kristin L.M.; Vance, Carroll P.

doi:10.1104/pp.84.2.501

Cited by 45 publications

(19 citation statements)

References 15 publications

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“…4, lanes 1 and 3) showed much less intense bands. These bands correspond to the 104-kD PEPC subunits in alfalfa and lupine root nodules (Miller et al, 1987). The PEPC activity of extracts used for immunoblot analysis was similar to that in Table II, with PEPC activity of proteoid roots being significantly greater than normal roots.…”

Section: Pepc Immunoblot Analysismentioning

confidence: 55%

“…Protein extracted from alfalfa nodules was used as a positive control for antibody detection. Miller et al (1987) demonstrated that this antibody cross-reacts with the 100-kD lupin nodule PEPC protein and the 100-kD alfalfa root PEPC protein.…”

Section: Sds-page and Western Blot Lmmunoanalysis For Pepcmentioning

confidence: 99%

“…Soluble protein (50 pg of protein for each tissue) from cell-free extracts was separated by SDS-PAGE and electrophoretically transferred to nitrocellulose. Rabbit polyclonal antibodies to alfalfa nodule PEPC were used to detect lupin root PEPC protein on westem blots (Miller et al, 1987). Protein extracted from alfalfa nodules was used as a positive control for antibody detection.…”

Section: Sds-page and Western Blot Lmmunoanalysis For Pepcmentioning

confidence: 99%

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Phosphorus Stress-Induced Proteoid Roots Show Altered Metabolism in Lupinus albus

1994

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Proteoid roots develop in Lupinus albus 1. in response to nutrient stress, especially P. Proteoid roots excrete citrate and thus increase the availability of P, Fe, and Mn in the rhizosphere. In an effort to understand citrate synthesis and organic acid metabolism in proteoid roots of lupin, we have evaluated i n vitro enzyme activities of citrate synthase (CS), malate dehydrogenase (MDH), and phosphoenolpyruvate carboxylase (PEPC) i n proteoid and normal roots of plants grown with or without P. Organic acid concentrations, respiration rates, and dark ''COz-labeling patterns were also determined. The in vitro specific activities of CS, MDH, and PEPC and in vivo dark "COZ fixation were higher i n proteoid roots compared to normal roots, particularly under P stress. Western blot analysis showed that PEPC enzyme protein was more highly expressed in -P proteoid roots compared to other tissues. The majority of the fixed 14C was found in organic acids, predominantly malate and citrate. A larger fraction of citrate was labeled in Pstressed proteoid roots compared to other root tissue. Respiration rates of proteoid roots were 31% less than those of normal roots. The data provide evidence for increased synthesis of citrate in proteoid roots compared to normal roots, particularly under P stress. A portion of the carbon for citrate synthesis is derived from nonautotrophic COz fixation via PEPC in proteoid roots.

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Section: Pepc Immunoblot Analysismentioning

confidence: 55%

Section: Sds-page and Western Blot Lmmunoanalysis For Pepcmentioning

confidence: 99%

Section: Sds-page and Western Blot Lmmunoanalysis For Pepcmentioning

confidence: 99%

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Phosphorus Stress-Induced Proteoid Roots Show Altered Metabolism in Lupinus albus

1994

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“…Alfalfa nodules are also reported to have Fd-GOGAT activity (A Suzuki, personal communication). Cross-reaction ofalfalfa nodule NADH-GOGAT antibodies with a polypeptide of comparable mol wt from other legume nodules is consistent with the cross-reaction ofPhaseolus nodule GS (5) and alfalfa nodule phosphoenol-pyruvate carboxylase (PEPC) antibodies (14) with other legume GS and PEPC polypeptides.…”

Section: Glutamate Synthase From Alfalfa Root Nodulesmentioning

confidence: 65%

“…In view of the lack of information on nodule GOGAT, the unavailability of antibodies to the NADH form of the enzyme, and the crucial role of GOGAT in nodule and plant N metabolism, the objectives of this research were to: (a) purify plant NADH-dependent GOGAT from alfalfa root nodules, (b) produce polyclonal antibodies to the enzyme, and (c) characterize the mol wt and some basic kinetic parameters of the enzyme. 400 Amol m-2 s-' as previously described (9,14,27). Before flowering, plants were gently uprooted and washed free of sand.…”

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confidence: 99%

Purification and Characterization of NADH-Glutamate Synthase from Alfalfa Root Nodules

Anderson¹,

Vance²,

Heichel³

et al. 1989

Plant Physiol.

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Glutamate synthase (GOGAT), a key enzyme in the pathway for the assimilation of symbiotically fixed dinitrogen (N2) into amino acids in alfalfa (Medicago sativa L.) root nodules, was purified and used to produce high titer polyclonal antibodies. Purification resulted in a 208-fold increase in specific activity to 13 micromole per minute per milligram of protein and an activity yield of 37%. Further purification to near homogeneity was achieved by fast protein liquid chromatography, but with substantial loss of activity. Enzymic activity was highly labile, losing 3% per hour even when substrates, stabilizers, and reducing agents were included in buffers. However, activity could be partially stabilized for up to 1 month by storing GOGAT at -800C in 50% glycerol. The subunit molecular weight of GOGAT was estimated at 200 ± 7 kilodaltons with a native molecular weight of 235 ± 16 kilodaltons, which suggested that GOGAT is a monomer of unusually high molecular weight. The pi was estimated to be 6.6. The Km values for glutamine, a-ketoglutarate, and NADH were 466, 33, and 4.2 micromolar, respectively. Antibodies were produced to NADH-GOGAT. Specificity of the antibodies was shown by immunotitration of GOGAT activity. Alfalfa nodule NADH-GO-GAT antibodies cross-reacted with polypeptides of a similar molecular weight in a number of legume species. Westem blots probed with anti-GOGAT showed that the high GOGAT activity of nodules as compared to roots was associated with increased levels of GOGAT polypeptides. Nodule NADH-GOGAT appeared to be highly expressed in effective nodules and little if any in other organs.

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How are nitrogen fixation rates regulated in legumes?

Schulze¹

2004

Z. Pflanzenernähr. Bodenk.

137

101

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± ZusammenfassungSymbiotic nitrogen fixation by the legume-Rhizobium symbiosis is a finely regulated process that involves significant carbon and energy metabolism by the plant. At present, there are three competing theories as to how the regulation of the process is achieved at the whole plant level. Theory one (carbon supply regulation) states that nitrogen fixation rates are regulated by either current nodule assimilate supply or through metabolism of assimilates into compounds that can be used by the bacteroids. A second theory (oxygen supply regulation) assumes that oxygen diffusion into the nodules is tightly regulated and is the principal regulatory factor for nitrogen fixation rates. A third theory (N-feedback regulation) suggests that a product of nitrogen fixation or assimilation exerts a feedback regulatory impact. The paper summarizes experimental data which support or reject the mentioned theories. Moreover, implications of nitrogen fixation under P stress and of phenomena connected with the argon or acetylene induced decline in nitrogenase activity for the understanding of nitrogen fixation regulation are discussed. It is concluded that we currently have no theory that explains all aspects and experimental results concerning the regulation of nitrogen fixation in legumes. With our growing ability to impact the process, e.g., by genetic engineering, this understanding of nitrogen fixation regulation has potential to be translated into agronomically sustainable benefits. Wie werden die Stickstofffixierungsraten bei Leguminosen reguliert?Die symbiontische N 2 -Fixierung bei Leguminosen ist ein fein regulierter Prozess, der einen massiven Substanzumsatz in der Pflanze beinhaltet. Gegenwärtig gibt es drei konkurrierende Theorien über die Art, wie die Regulation des Prozesses auf Ganzpflanzenbasis erreicht wird. Eine Theorie (Regulation über die Kohlenstoffversorgung) geht von einer Abhängigkeit bzw. Regulation der N 2 -Fixierungsraten entweder durch die Assimilatversorgung der Knöllchen oder von einer Limitierung des nötigen metabolischen Umsatzes der Assimilate in für den Bakteroiden verwertbare Verbindungen aus. In der zweiten Theorie (Regulation über die Sauerstoffversorgung) wird angenommen, dass die Sauerstoffdiffusion in das Knöllchen einer strengen Regulation unterliegt und dies der prinzipielle Regulationsfaktor für die N 2 -Fixierungsraten ist. Die dritte Theorie (ªN-feedbackº-Regulation) besagt, dass ein bisher nicht zweifelsfrei identifiziertes Produkt der Stickstoffixierung oder -assimilation eine ªfeedbackª-Wirkung auf die N 2 -Fixierung ausübt. Der vorliegende Artikel fasst die bis heute gesammelten experimentellen Daten zusammen, die für oder gegen die genannten Theorien sprechen. In diesem Zusammenhang wird diskutiert, auf welche Weise die Regulation der N 2 -Fixierung unter P-Mangelbedingungen erfolgt. Schlieûlich wird diskutiert, ob die Klärung der Ursache der Azetylen-bzw. Argon-induzierten Abnahme der Nitrogenaseaktivität zum Verständnis der Regulation der N 2 -Fixierung beitragen kan...

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Alfalfa Root Nodule Carbon Dioxide Fixation

Cited by 45 publications

References 15 publications

Phosphorus Stress-Induced Proteoid Roots Show Altered Metabolism in Lupinus albus

Phosphorus Stress-Induced Proteoid Roots Show Altered Metabolism in Lupinus albus

Purification and Characterization of NADH-Glutamate Synthase from Alfalfa Root Nodules

How are nitrogen fixation rates regulated in legumes?

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