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1.
Nitrate addition to nitrate-limited cultures of Selenastrum minutum Naeg. Collins (Chlorophyta) resulted in a 70% suppression of photosynthetic carbon fixation. In 14CO2 pulse/chase experiments nitrate resupply increased radiolabel incorporation into amino and organic acids and decreased radiolabel incorporation into insoluble material. Nitrate resupply increased the concentration of phosphoenolpyruvate and increased the radiolabeling of phosphoenolpyruvate, pyruvate and tricarboxylic acid cycle intermediates, notably citrate, fumarate, and malate. Furthermore, nitrate also increased the pool sizes and radiolabeling of most amino acids, with alanine, aspartate, glutamate, and glutamine showing the largest changes. Nitrate resupply increased the proportion of radiolabel in the C-4 position of malate and increased the ratios of radiolabel in aspartate to phosphoenolpyruvate and in pyruvate to phosphoenolpyruvate, indicative of increased phosphoenolpyruvate carboxylase and pyruvate kinase activities. Analysis of these data showed that the rate of carbon flow through glutamate (10.6 μmoles glutamate per milligram chlorophyll per hour) and the rate of net glutamate production (7.9 μmoles glutamate per milligram chlorophyll per hour) were both greater than the maximum rate of carbon export from the Calvin cycle which could be maintained during steady state photosynthesis. These results are consistent with the hypothesis that nitrogen resupply to nitrogen-limited microalgae results in a transient suppression of photosynthetic carbon fixation due, in part, to the severity of competition for carbon skeletons between the Calvin cycle and nitrogen assimilation (IR Elrifi, DH Turpin 1986 Plant Physiol 81: 273-279).  相似文献   

2.
Nobel PS 《Plant physiology》1967,42(10):1389-1394
The main features of the procedure developed for rapid chloroplast isolation are: 1) gentle grinding of the plant material in a special nylon bag which retains nearly all whole cells and large debris, 2) osmoticum concentration chosen on the basis of the measured endogenous photophosphorylation, 3) a single, brief, low-speed centrifugation, 4) pellet resuspension by means of a vortex mixer, and 5) a total elapsed time from harvesting the plants to the obtaining of a resuspended chloroplast pellet of only 2 minutes. The usual isolation medium consists of an osmoticum (0.2 m sucrose) and a buffer (0.02 m N-tris-(hydroxymethyl) methyl-2-aminoethanesulfonate-NaOH, pH 7.9). In addition to these, the incubation medium contains only 200 μm ADP and 200 μm phosphate. Photophosphorylation rates of 24 μmoles ATP formed per mg chlorophyll per hour are consistently obtained using chloroplasts isolated from peas (Pisum sativum var. Laxton's Superb). The rate of endogenous photophosphorylation is maximal when the isolation and incubation media have an osmolarity of about 0.19 made up either with sucrose or with NaCl. The high rates and ease of measurement of endogenous photophosphorylation may facilitate the study of certain soluble components of chloroplasts as well as the general state of the photosynthetic ability of the plant.  相似文献   

3.
Autocatalysis in a reconstituted chloroplast system   总被引:2,自引:2,他引:0       下载免费PDF全文
In whole plants and intact chloroplasts, photosynthesis does not reach its full rate immediately upon illumination but only after a lag which is believed to reflect an autocatalytic increase in the concentration of carbon cycle intermediates. Autocatalysis has now been observed in a reconstituted system containing envelope-free chloroplasts augmented with ferredoxin and other stromal proteins but only catalytic amounts of ATP and NADP. With ribose 5-phosphate as substrate, the CO2 dependent O2 evolution recorded for such mixtures implies rates of “endogenous” or ferredoxin-dependent photophosphorylation as high as 360 μmoles of orthophosphate esterified mg−1 chlorophyll hr−1.  相似文献   

4.
The assimilation of nitrite leading to de novo synthesis of amino nitrogen in a chloroplast-enriched fraction isolated from freshly harvested young spinach (Spinacia oleracea L.) leaves was demonstrated. The preparations showed approximately 55% intact chloroplasts as determined by light scattering properties and fixed CO2 at rates of approximately 100 μmoles hr−1 mg chlorophyll−1.  相似文献   

5.
Yu J  Woo KC 《Plant physiology》1988,88(4):1048-1054
The transport of l-[14C]glutamine in oat (Avena sativa L.) and spinach (Spinacia oleracea L.) chloroplasts was studied by a conventional single-layer and a newly developed stable double-layer silicone oil filtering system. [14C]Glutamine was actively transported into oat chloroplasts against a concentration gradient. Metabolite uptake was greatly affected by the endogenous dicarboxylate pools, which could be easily changed by preloading the chloroplast with specific exogenous substrate. Glutamine uptake was decreased by 44 to 75% in oat chloroplasts preloaded with malate, 2-oxoglutarate (2-OG), and aspartate, but increased by 52% in chloroplasts preloaded with l-glutamate. On the other hand, the uptake of the other four dicarboxylates was decreased by 47 to 79% in chloroplasts preloaded with glutamine. In glutamine-preloaded chloroplasts the uptake of glutamine was inhibited only by l-glutamate. The observed inhibition by l-glutamate was competitive with an apparent Ki value of 32.1 millimolar in oat and 6.7 millimolar in spinach chloroplasts. This study indicates that there are two components involved in glutamine transport in chloroplasts. The major component was mediated via a specific glutamine translocator. It was specific for glutamine and did not transport other dicarboxylates except l-glutamate. A K0.5 value of 1.25 millimolar and Vmax of 45.5 micromoles per milligram of chlorophyll per hour were determined for the glutamine translocator in oat chloroplasts. The respective values were 1.0 millimolar and 16.7 micromoles per milligram of chlorophyll per hour in spinach chloroplasts. A three translocator model, involving the glutamine, dicarboxylate, and 2-OG translocators, is proposed for the reassimilation of photorespiratory NH3 in chloroplasts of C3 species. In this three-translocator model the additional transport of glutamine into the chloroplast is coupled to the export of glutamate via the glutamine translocator. This is an extension of the two-translocator model, involving the dicarboxylate and 2-OG translocators, proposed for spinach chloroplasts, (KC Woo, UI Flügge, HW Heldt 1987 Plant Physiol 84: 624-632).  相似文献   

6.
Haploid callus cells of tobacco (Nicotiana tabacum) were grown photoautotrophically on a solid agar medium in the absence of sucrose in Petri plates in an atmosphere of 1% or 3% CO2 in air. The averages of dry weight increases for four to five consecutive passages were 2.3- to 3.6-fold per 3-week passage for different subclones. Photosynthetic 14CO2 assimilation was maximum at about 1% CO2 with half-maximal rates obtained at 0.2% CO2. At saturating CO2 concentration the average rate of CO2 fixation was about 5 μmole per gram fresh weight per hour or about 125 μmole per mg of chlorophyll per hour.  相似文献   

7.
Chen C  Gibbs M 《Plant physiology》1992,98(2):535-539
The reductive carboxylic acid cycle, the autotrophic pathway of CO2 assimilation in prokaryotes (photosynthetic and nonphotosynthetic autotrophic bacteria), was investigated in Chlamydomonas reinhardtii F-60, an algal mutant lacking a complete photosynthetic carbon reduction pathway (C3) due to a deficiency in phosphoribulokinase. Evidence was obtained consistent with the presence of the reductive carboxylic acid cycle in F-60. This conclusion is based on the fact that: (a) acetate approximately doubled CO2 fixation in whole cells (4 micromoles per milligram chlorophyll per hour) and in chloroplasts (32 nanomoles per milligram chlorophyll per hour); and (b) pyruvate synthase, α-ketoglutarate synthase, and ATP-citrate lyase, three indicators of the cycle, were found in cell-free extracts.  相似文献   

8.
Terry N 《Plant physiology》1976,57(4):477-479
Effects of sulfur on photosynthesis in sugar beets (Beta vulgaris L. cv. F58-554H1) were studied by inducing sulfur deficiency and determining changes in the photosynthesis of whole attached leaves and of isolated chloroplasts. The rates of photosynthetic CO2 uptake by intact leaves, photoreduction of ferricyanide, cyclic and noncyclic photophosphorylation of isolated chloroplasts, and the rate of CO2 assimilation by ribulose diphosphate carboxylase, decreased with decrease in total leaf sulfur from 2500 to about 500 μg g−1 dry weight. Sulfur deficiency reduced photosynthesis through an effect on chlorophyll content, which decreased linearly with leaf sulfur, and by decreasing the rate of photosynthesis per unit chlorophyll. There was only a small effect of sulfur deficiency on stomatal diffusion resistance to CO2 until leaf sulfur decreased below 1000 μg g−1 when stomatal resistance became a more significant proportion of the total diffusion resistance to CO2. Light respiration rates were positively correlated with photosynthesis rates and dark respiration was unchanged as leaf sulfur concentrations declined.  相似文献   

9.
Chloroplasts were prepared from peas (Pisum sativum) in glucose-phosphate medium. In the presence of dl-glyceraldehyde, they catalyzed nitrite-dependent O2 evolution (mean of 13 preparations, 17.5 μmole per mg chlorophyll per hour, sd 3.64). The optimum concentration of nitrite was 0.5 mm; 0.12 mm nitrite supported Vmax/2. The reaction was accompanied by the consumption of nitrite; 55 to 80% of the nitrite-N consumed was recovered as ammonia. In short experiments (less than 10 minutes) the O2 to nitrite ratio approached 1.5, but thereafter decreased. There was no nitrite-dependent O2 evolution with chloroplasts from plants grown without added nitrate but such chloroplasts could assimilate ammonia at about the usual rate. The results are consistent with the reduction of nitrite to ammonia involving nitrate-induced nitrite reductase and a reductant generated by the chloroplast electron transport chain.  相似文献   

10.
Carbon dioxide fixation in isolated kalanchoe chloroplasts   总被引:2,自引:2,他引:0       下载免费PDF全文
Levi C  Gibbs M 《Plant physiology》1975,56(1):164-166
Chloroplasts isolated from Kalanchoe diagremontiana leaves were capable of photosynthesizing at a rate of 5.4 μmoles of CO2 per milligram of chlorophyll per hour. The dark rate of fixation was about 1% of the light rate. A high photosynthetic rate was associated with low starch content of the leaves. Ribose 5-phosphate, fructose 1,6-diphosphate, and dithiothreitol stimulated fixation, whereas phosphoenolpyruvate and azide were inhibitors. The products of CO2 fixation were primarily those of the photosynthetic carbon reduction cycle.  相似文献   

11.
The drought-resistant cyanobacteria Phormidium autumnale, strain LPP4, and a Chroococcidiopsis sp. accumulated trehalose, sucrose, and both trehalose and sucrose, respectively, in response to matric water stress. Accumulated sugar concentrations reached values of up to 6.2 μg of trehalose per μg of chlorophyll in P. autumnale, 6.9 μg of sucrose per μg of chlorophyll in LPP4, and 4.1 μg of sucrose and 3.2 μg of trehalose per μg of chlorophyll in the Chroococcidiopsis sp. The same sugars were accumulated by these cyanobacteria in similar concentrations under osmotic water stress. Cyanobacteria that did not show drought resistance (Plectonema boryanum and Synechococcus strain PCC 7942) did not accumulate significant amounts of sugars when matric water stress was applied.  相似文献   

12.
Isolated maize bundle sheath chloroplasts showed substantial rates of noncyclic photophosphorylation. A typical rate of phosphorylation coupled to whole-chain electron transport (methylviologen or ferricyanide as acceptor) was 60 μmol per hour per milligram chlorophyll) with a coupling efficiency (P/e2) of 0.6. Partial electron transport reactions driven by photosystem I or II supported phosphorylation with P/e2 values of 0.2 to 0.3. Thus, two sites of phosphorylation seem to be associated with the photosynthetic chain in much the same way as in spinach chloroplasts.  相似文献   

13.
Fragments of bundle sheath strands, free of mesophyll cells and showing a chlorophyll a/b ratio of 6.0 to 6.6 were prepared from Zea mays by a mechanical method. They were unable to photoreduce ferricyanide but were able to photoreduce the membrane-permeant 2,5-dimethylquinone at a rate of 250 to 420 microequivalents per hour per mg chlorophyll (μeq/hr · mg Chl) at 21 C. In the presence of the catalase inhibitor KCN, methylviologen catalyzed a Mehler reaction at a rate of 120 to 180 μeq/hr · mg Chl. This was increased to 200 to 350 μeq/hr · mg Chl when the uncoupler methylamine was added. The rate of endogenous pseudocyclic electron flow, detected as a Mehler reaction, was also considerable (100 to 150 μeq/hr · mg Chl with methylamine). Diaminodurene supported a high rate of photosystem I-mediated electron flow to methylviologen (400 to 750 μeq/hr · mg Chl).  相似文献   

14.
Enzymic and substrate basis for the anaplerotic step in guard cells   总被引:4,自引:4,他引:0  
From the maximum rate of malate accumulation in Vicia faba L. guard cells during stomatal opening the maximum rate of organic anion synthesis is calculated to be 200 millimoles per kilogram dry weight per hour. A minimum estimate for the phosphoenolpyruvate (PEP) carboxylase-catalyzed reaction in guard cells is 650 millimoles per kilogram dry weight per hour which is significantly higher than in any other leaf tissue. The apparent Kmpep of the guard cell enzyme is 60 μm at pH 8.7, but is probably higher at lower pH. The concentration of PEP in guard cells was 270μm (=2.2 × 10−15 moles/guard cell pair) during anion synthesis. These results support the possibility that the carboxylation of PEP is the anaplerotic step in guard cells.  相似文献   

15.
Inhibition of chloroplast reactions with phenylmercuric acetate   总被引:1,自引:1,他引:0       下载免费PDF全文
Phenylmercuric acetate is a selective inhibitor of the photosynthetic activities of isolated spinach (Spinacia oleracea) chloroplasts. At 5 μm concentration of phenylmercuric acetate, photophosphorylation is inhibited. At 33 μm phenylmercuric acetate, ferredoxin is inactivated. Ferredoxin-NADP oxidoreductase is 50% inhibited at 100 μm phenylmercuric acetate. Photosystem II reactions are 50% inhibited at 150 μm phenylmercuric acetate and very much higher cooncentrations—500 μm—are needed to approach complete inhibition. Phenylmercuric acetate inhibition of photosystem II appears to be selective, blocking a site between the 3-(3,4-dichlorophenyl)-1,1-dimethyl urea sensitive site and the site inactivated by high concentrations of tris buffer.  相似文献   

16.
Nodulated bean (Phaseolus vulgaris) plants were grown for 17 days after infection in normal (0.02%) CO2 and from day 8 to 17 in high (0.1%) CO2 in order to increase nitrogen fixation and define how nodule glutamine synthetase (GS) isoforms are regulated by the ammonia derived from the bacteroid. Nitrogenase activity was detected by day 10, and by day 17 activity was over twofold higher in 0.1% of CO2 compared with plants grown in 0.02% CO2 and inoculated with Rhizobium wild-type strain CE3. Likewise, plant fresh weight increased in response to increased CO2, particularly in plants inoculated with the Rhizobium phaseoli mutant strain CFN037. Glutamine synthetase specific activity increased 2.5- to 6.5-fold from day 11 to 17. However, increased CO2 did not appear to have an effect on GS specific activity. Analysis of the nodule GS polypeptide composition revealed that the γ polypeptide was significantly reduced in response to high CO2, whereas the β polypeptide was not affected. The significance of this result in relation to the regulation of GS isoforms and their role in the assimilation of ammonia in the nodule is discussed in this paper.  相似文献   

17.
The establishment and maintenance of high rates of photosynthetic CO2 incorporation in mesophyll cells of Papaver somniferum (opium poppy) depend on a regime of dark and light periods immediately following isolation, as well as carefully adjusted conditions of isolation. Analysis of the incorporation pattern of 14CO2 by the isolated cells indicates an initial “stress-response” period of approximately 20 hours characterized by increased respiratory-type metabolism and diminished photosynthesis. Under the favorable regime, this period is followed by rapid recovery and the reinstatement of a metabolic state strikingly similar to that of intact leaves in which the initial rate of CO2 incorporation is between 110 and 175 μmoles CO2 fixed per mg chlorophyll per hour. The photosynthetic viability of these cells can be maintained for up to 80 hours.  相似文献   

18.
The protein components of the 2-nitrotoluene (2NT) and nitrobenzene dioxygenase enzyme systems from Acidovorax sp. strain JS42 and Comamonas sp. strain JS765, respectively, were purified and characterized. These enzymes catalyze the initial step in the degradation of 2-nitrotoluene and nitrobenzene. The identical shared reductase and ferredoxin components were monomers of 35 and 11.5 kDa, respectively. The reductase component contained 1.86 g-atoms iron, 2.01 g-atoms sulfur, and one molecule of flavin adenine dinucleotide per monomer. Spectral properties of the reductase indicated the presence of a plant-type [2Fe-2S] center and a flavin. The reductase catalyzed the reduction of cytochrome c, ferricyanide, and 2,6-dichlorophenol indophenol. The ferredoxin contained 2.20 g-atoms iron and 1.99 g-atoms sulfur per monomer and had spectral properties indicative of a Rieske [2Fe-2S] center. The ferredoxin component could be effectively replaced by the ferredoxin from the Pseudomonas sp. strain NCIB 9816-4 naphthalene dioxygenase system but not by that from the Burkholderia sp. strain LB400 biphenyl or Pseudomonas putida F1 toluene dioxygenase system. The oxygenases from the 2-nitrotoluene and nitrobenzene dioxygenase systems each had spectral properties indicating the presence of a Rieske [2Fe-2S] center, and the subunit composition of each oxygenase was an α3β3 hexamer. The apparent Km of 2-nitrotoluene dioxygenase for 2NT was 20 μM, and that for naphthalene was 121 μM. The specificity constants were 7.0 μM−1 min−1 for 2NT and 1.2 μM−1 min−1 for naphthalene, indicating that the enzyme is more efficient with 2NT as a substrate. Diffraction-quality crystals of the two oxygenases were obtained.  相似文献   

19.
Bundle sheath chloroplasts have been isolated from Zea mays leaves by a procedure involving enzymic digestion of mechanically prepared strands of bundle sheath cells followed by gentle breakage and filtration. The resulting crude chloroplast preparation was enriched by Percoll density layer centrifugation to yield intact chloroplasts (about 20 micrograms chlorophyll per 10-gram leaf tissue) with high metabolic activities. Based on activities of marker enzymes in the chloroplast and bundle sheath cell extracts, the chloroplasts were essentially free of contamination by other organelles and cytoplasmic material, and were generally about 70% intact. Chlorophyll a/b ratios were high (about 10). With appropriate substrates these chloroplasts displayed high rates of malate decarboxylation, measured as pyruvate formation, and CO2 assimilation (maximum rates approximately 5 and 3 micromoles per minute per milligram chlorophyll, respectively). These activities were light dependent, linear for at least 20 minutes at 30°C, and displayed highest rates at pH 8.0. High metabolic rates were dependent on addition of an exogenous source of carbon to the photosynthetic carbon reduction cycle (3-phosphoglycerate or dihydroxyacetone phosphate) and a nucleotide (ATP, ADP, or AMP), as well as aspartate. Generally, neither malate decarboxylation nor CO2 assimilation occurred substantially in the absence of the other activity indicating a close relationship between these processes. Presumably, NADPH required for the photosynthetic carbon reduction cycle is largely supplied during the decarboxylation of malate by NADP-malic enzyme. The results are discussed in relation to the role of bundle sheath chloroplasts in C4 photosynthesis by species of the NADP-malic enzyme type.  相似文献   

20.
Levels of nitrate reductase activity (EC 1.9.6.1.) as high as 11 μmoles nitrite produced/hour gram fresh weight were found in barley (Hordeum vulgare cv. Compana) roots grown under low oxygen conditions. Roots of plants given identical treatment under sterile conditions did not develop the high levels of nitrate reductase activity. The results suggest that the buildup of particulate, reduced viologen-utilizing nitrate reductase reported in barley roots may be caused by bacterial contamination. The nitrate reductase activity in roots grown under low oxygen conditions was not specific for reduced nicotinamide adenine dinucleotide like the assimilatory nitrate reductase (EC 1.6.6.1.) normally found in aerated plant roots.  相似文献   

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