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1.
Methionine sulfoximine induced release of ammonia from illuminated cells of Ankistrodesmus braunii (Naegeli) Brunnth, in normal air, but less in air enriched to 3% CO 2. In normal air, methionine sulfoximine also induced glycolate release. Addition of either glutamate, glycine, or serine suppressed glycolate release, whereas glutamate and glycine at the same time stimulated ammonia release. The results indicate that inhibition of glutamine synthetase and thereby inhibition of photorespiratory nitrogen cycling restricts the sink capacity for glycolate in the photorespiratory carbon cycle. An external supply of glutamate, glycine, or serine seems to stimulate glyoxylate transamination and thus partly restores the sink capacity. Calculations of total glycolate formation rates in air from glycolate and ammonia release rates in the presence of methionine sulfoximine and glutamate revealed values of approximately 20 micromoles glycolate per milligram chlorophyll per hour on the average. Similar calculations led to an estimated rate of photorespiratory ammonia release in air, in the absence of methionine sulfoximine, of about 10 micromoles per milligram chlorophyll per hour on the average, a value comparable to the primary nitrogen assimilation rate of 8 micromoles per milligram chlorophyll per hour. 相似文献
2.
Photorespiration in Chlorella pyrenoidosa Chick. was assayed by measuring 18O-labeled intermediates of the glycolate pathway. Glycolate, glycine, serine, and excreted glycolate were isolated and analyzed on a gas chromatograph/mass spectrometer to determine isotopic enrichment. Rates of glycolate synthesis were determined from 18O-labeling kinetics of the intermediates, pool sizes, derived rate equations, and nonlinear regression techniques. Glycolate synthesis was higher in high CO 2-grown cells than in air-grown cells when both were assayed under the same O 2 and CO 2 concentrations. Synthesis of glycolate, for both types of cells, was stimulated by high O 2 levels and inhibited by high CO 2 levels. Glycolate synthesis in 1.5% CO 2-grown Chlorella, when exposed to a 0.035% CO 2 atmosphere, increased from about 41 to 86 nanomoles per milligram chlorophyll per minute when the O 2 concentration was increased from 21% to 40%. Glycolate synthesis in air-grown cells increased from 2 to 6 nanomoles per milligram chlorophyll per minute under the same gas levels. Synthesis was undetectable when either the O 2 concentration was lowered to 2% or the CO 2 concentration was raised to 1.5%. Glycolate excretion was also sensitive to O 2 and CO 2 concentrations in 1.5% CO 2-grown cells and the glycolate that was excreted was 18O-labeled. Air-grown cells did not excrete glycolate under any experimental condition. Indirect evidence indicated that glycolate may be excreted as a lactone in Chlorella. Photorespiratory 18O-labeling kinetics were determined for Pavlova lutheri, which unlike Chlorella and higher plants did not directly synthesize glycine and serine from glycolate. This alga did excrete a significant proportion of newly synthesized glycolate into the media. 相似文献
3.
Glycolate and ammonia excretion plus oxygen exchanges were measured in the light in l-methionine- dl-sulfoximine treated air-grown Chlamydomonas reinhardii. At saturating CO 2 (between 600 and 700 microliters per liter CO 2) neither glycolate nor ammonia were excreted, whereas at the CO 2 compensation concentration (<10 microliters per liter CO 2) treated algae excreted both glycolate and ammonia at rates of 37 and 59 nanomoles per minute per milligram chlorophyll, respectively. From the excretion values we calculate the amount of O 2 consumed through the glycolate pathway. The calculated value was not significantly different from the component of O 2 uptake sensitive to CO 2 obtained from the difference between O 2 uptake of the CO 2 compensation point and at saturating CO 2. This component was about 40% of stationary O 2 uptake measured at the CO 2 compensation point. From these data we conclude that glyoxylate decarboxylation in air-grown Chlamydomonas represents a minor pathway of metabolism even in conditions where amino donors are deficient and that processes other than glycolate pathway are responsible for the O 2 uptake insensitive to CO 2. 相似文献
4.
Aerobic and anaerobic chloroplastic respiration was monitored by measuring 14CO 2 evolution from [ 14C]glucose in the darkened spinach ( Spinacia oleracea) chloroplast and by estimating the conversion of fructose 1,6-bisphosphate to glycerate 3-phosphate in the darkened spinach chloroplast in air with O 2 or in N 2 with nitrite or oxaloacetate as electron acceptors. The pathway of 14CO 2 evolution from labeled glucose in the absence and presence of the inhibitors iodoacetamide and glycolate 2-phosphate under air or N 2 were those expected from the oxidative pentose phosphate cycle and glycolysis. Of the electron acceptors, O 2 was the best (2.4 nanomoles CO 2 per milligram chlorophyll per hour), followed by nitrite and oxaloacetate. With respect to glycerate 3-phosphate formation from fructose 1,6-bisphosphate, methylene blue increased the aerobic rate from 3.7 to 5.4 micromoles per milligram chlorophyll per hour. A rate of 4.8 micromoles per milligram chlorophyll per hour was observed under N 2 with nitrite and oxaloacetate. 相似文献
5.
Cells of Dunaliella tertiolecta from the log phase of growth were broken by rapid extrusion at low pressure through a Yeda press and the chloroplasts were isolated by centrifugation through a Percoll gradient. Osmolarity of the growth media, the suspending media, and the Percoll gradient was kept identical to minimize change in chloroplast volume and mitochondrial entrapment. The isolated intact chloroplasts were obtained in a 30 to 50% yield based on chlorophyll and were stable to washing with buffered medium. Isolated chloroplast yield and purity was dependent on cell culture condition; a cycle of 16 hours light and 8 hours dark with continuous high CO 2 was optimum. Isolated chloroplasts were about 90% intact by microscopic examination, ferricyanide-dependent O 2 evolution, and the distribution of four stromal enzymes. Enzymes associated with glycolate metabolism were not in the chloroplast fraction. The isolated chloroplasts with 10 millimolar bicarbonate evolved 24 micromoles of O 2 and fixed 21 micromoles of CO 2 per hour per milligram of chlorophyll, which rates were about one-third of those by whole cells. The inhibition of oxygen evolution by 10 millimolar phosphate was reversed by P-glycerate. Whole chloroplasts were also isolated from cells adapted to low CO 2 in air for 24 hours. On low CO 2 the cells excreted more gelatinous material, which had to be removed with additional washing of the cells, before it was possible to obtain good chloroplast preparations. 相似文献
6.
A system has been developed for the isolation of photosynthetically active chloroplasts from leaves of Populus deltoides. A high proportion of the chloroplasts appeared intact. The maximum rates of different photosynthetic processes were as follows: CO 2 fixation 3.5 micromoles per milligram chlorophyll per hour, noncyclic ATP synthesis 10 micromoles per milligram chlorophyll per hour, and cyclic ATP synthesis 300 micromoles per milligram chlorophyll per hour. 相似文献
7.
A closed system consisting of an assimilation chamber furnished with a membrane inlet from the liquid phase connected to a mass spectrometer was used to measure O 2 evolution and uptake by Chlamydomonas reinhardtii cells grown in ambient (0.034% CO 2) or CO 2-enriched (5% CO 2) air. At pH = 6.9, 28°C and concentrations of dissolved inorganic carbon (DIC) saturating for photosynthesis, O 2 uptake in the light (U o) equaled O 2 production (E o) at the light compensation point (15 micromoles photons per square meter per second). E o and U o increased with increasing photon fluence rate (PFR) but were not rate saturated at 600 micromoles photons per square meter per second, while net O 2 exchange reached a saturation level near 500 micromoles photons per square meter per second which was nearly the same for both, CO 2-grown and air-grown cells. Comparison of the U o/E o ratios between air-grown and CO 2-grown C. reinhardtii showed higher values for air-grown cells at light intensities higher than light compensation. For both, air-grown and CO 2-grown algae the rates of mitochondrial O 2 uptake in the dark measured immediately before and 5 minutes after illumination were much lower than U o at PFR saturating for net photosynthesis. We conclude that noncyclic electron flow from water to NADP + and pseudocyclic electron flow via photosystem I to O 2 both significantly contribute to O 2 exchange in the light. In contrast, mitochondrial respiration and photosynthetic carbon oxidation cycle are regarded as minor O 2 consuming reactions in the light in both, air-grown and CO 2-grown cells. It is suggested that the “extra” O 2 uptake by air-grown algae provides ATP required for the energy dependent CO 2/HCO 3− concentrating mechanism known to be present in these cells. 相似文献
8.
The reductive carboxylic acid cycle, the autotrophic pathway of CO 2 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 (C 3) 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 CO 2 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
9.
Diffusion of inorganic carbon into isolated bundle sheath cells from a variety of C 4 species was characterized by coupling inward diffusion of CO 2 to photosynthetic carbon assimilation. The average permeability coefficient for CO 2 ( PCO2) for five representatives from the three decarboxylation types was approximately 20 micromoles per minute per milligram chlorophyll per millimolar, on a leaf chlorophyll basis. The average value for the NAD-ME species Panicum miliaceum (10 determinations) was 26 with a standard deviation of 6 micromoles per minute per milligram chlorophyll per millimolar, on a leaf chlorophyll basis. A PCO2 of at least 500 micromoles per minute per milligram chlorophyll per millimolar was determined for cells isolated from the C 3 plant Xanthium strumarium. It is concluded that bundle sheath cells are one to two orders of magnitude less permeable to CO 2 than C 3 photosynthetic cells. These data also suggest that CO 2 diffusion in bundle sheath cells may be made up of two components, one involving an apoplastic path and the other a symplastic (plasmodesmatal) path, each contributing approximately equally. 相似文献
10.
The in vitro specific activity of ribulose-1,5-bisphosphate carboxylase (RuBPCase) (micromoles CO 2 fixed per minute per milligram enzyme) from a number of C 3 and C 4 species and one green alga were measured. RuBPCases from species which utilize the C 4 pathway have a specific activity ~2-fold higher than those from C 3 species. RuBPCase from Chlamydomonas reinhardtii has a specific activity similar to the C 4 enzyme. Higher specific activity forms of RuBPCase are associated with a decreased enzyme affinity for CO 2 (increased K m[CO 2]). A small but significant difference in the specific activity of RuBPCase from two C 4 decarboxylation types was also observed. The relationship between enzymic properties and the presence or absence of a CO 2 concentrating mechanism is discussed. 相似文献
11.
Aminooxyacetate (1 millimolar) did not inhibit photosynthetic 14CO 2 fixation by Chlamydomonas reinhardtii Dangeard, (−) strain (N.90) but greatly stimulated the biosynthesis and excretion of glycolate. Similar results were obtained from cells grown with 5% CO 2 or low CO 2 (air). After 2 minutes with air-grown cells, [ 14C]glycolate increased from 0.3% of the total 14C fixed by the control to 11.7% in the presence of aminooxyacetate and after 10 minutes from 3.8% to 41.1%. Ammonium nitrate (0.2 millimolar) in the media blocked the aminooxyacetate stimulation of glycolate excretion. Chromatographic analyses of the labeled products in the cells and supernatant media indicated that aminooxyacetate also completely inhibited the labeling of alanine while some pyruvate accumulated and was excreted. A high percentage (35%) of initial 14CO 2 fixation was into C 4 acids. Initial products of 14CO 2 fixation included phosphate esters as well as malate, aspartate, and glutamate in treated or untreated cells. Lactate was also a major early product of photosynthesis, and its labeling was reduced by aminooxyacetate. Inasmuch as lactate was not excreted, glycolate excretion seemed to be specific. When photosynthesis was inhibited by 3-(3,4-dichlorophenyl)-1,1-dimethylurea, labeled organic and amino acids but not phosphate esters were lost from the cells. Aminooxyacetate did not inhibit the enzymes associated with glycolate synthesis from ribulose bisphosphate. 相似文献
12.
The claim that Chlorella sp. (CCAP 211/8p), sometimes referred to as C. fusca, Shihira and Krauss, does not excrete glycolate has been reexamined. Chlorella sp. grown on 5% CO2in air, excreted glycolate when incubated in light in 10 mM bicarbonate. Excretion ceased 30–60 min after transfer of the cells to air and no excretion could be detected with air-grown cells or with cells grown on 5% CO2in media buffered at pH 8.0. Incubation with 10 mM isonicotinyl hydrazide, a glycolate pathway inhibitor, caused excretion in air-grown cells and stimulated excretion in CO2-grown cells indicating that both the rate of glycolate synthesis and metabolism is higher in CO2grown cells than in air-grown cells. Enhanced glycolate synthesis and excretion in CO2-grown cells is correlated with law photosynthetic rate in 10 mM bicarbonate, and the photosynthetic rate of these cells doubles over a period of 2–2.5 h after initial transfer from high CO2to bicarbonate. This correlation of photosynthetic induction with cessation of glycolate excretion is similar to that reported in a bluegreen alga and thought to occur in other green algae. These results indicate that glycolate excretion and its regulation in this species of Chlorella is not different from that in other algae. 相似文献
13.
Young expanding spinach leaves exposed to 14CO 2 under physiological conditions for up to 20 minutes assimilated CO 2 into lipids at a mean rate of 7.6 micromoles per milligram chlorophyll per hour following a lag period of 5 minutes. Label entered into all parts of the lipid molecule and only 28% of the 14C fixed into lipids was found in the fatty acid moieties, i.e. fatty acids were synthesized from CO 2in vivo at a mean rate of 2.1 micromoles per milligram chlorophyll per hour. Intact spinach chloroplasts isolated from these leaves incorporated H 14CO 3 into fatty acids at a maximal rate of 0.6 micromole per milligram chlorophyll per hour, but were unable to synthesize either the polar moieties of their lipids or polyunsaturated fatty acids. Since isolated chloroplasts will only synthesize fatty acids at rates similar to the one obtained with intact leaves in vivo if acetate is used as a precursor, it is suggested that acetate derived from leaf mitochondria is the physiological fatty acid precursor. 相似文献
14.
Mechanically isolated soybean leaf cells metabolized added glycolate by two mechanisms, the direct oxidation of glyoxylate and the decarboxylation of glycine. The rate of glyoxylate oxidation was dependent on the cellular glyoxylate concentration and was linear between 0.58 and 2.66 micromoles glyoxylate per milligram chlorophyll. The rate extrapolated to zero at a concentration of zero. The concentration and, therefore, the rate of oxidation of glyoxylate could be decreased by adding glutamate or serine to the cells. These substrates were amino donors for the transamination of glyoxylate to glycine. In the presence of these amino acids more CO 2 was released from added glycolate via the glycine decarboxylation reaction and less by the direct oxidation of glyoxylate. 相似文献
15.
Mesophyll cells and bundle sheath strands were isolated rapidly from leaves of the C 4 species Digitaria pentzii Stent. (slenderstem digitgrass) by a chopping and differential filtration technique. Rates of CO 2 fixation in the light by mesophyll and bundle sheath cells without added exogenous substrates were 6.3 and 54.2 micromoles of CO 2 per milligram of chlorophyll per hour, respectively. The addition of pyruvate or phosphoenolpyruvate to the mesophyll cells increased the rates to 15.2 and 824.6 micromoles of CO 2 per milligram of chlorophyll per hour, respectively. The addition of ribose 5-phosphate increased the rate for bundle sheath cells to 106.8 micromoles of CO 2 per milligram of chlorophyll per hour. These rates are comparable to those reported for cells isolated by other methods. The Km(HCO 3−) for mesophyll cells was 0.9 m m; for bundle sheath cells it was 1.3 m m at low, and 40 m m at higher HCO 3− concentrations. After 2 hours of photosynthesis by mesophyll cells in 14CO 2 and phosphoenolpyruvate, 88% of the incorporated 14C was found in organic acids and 0.8% in carbohydrates; for bundle sheath cells incubated in ribose 5-phosphate and ATP, more than 58% of incorporated 14C was found in carbohydrates, mainly starch, and 32% in organic acids. These findings, together with the stimulation of CO 2 fixation by phosphoenolpyruvate for mesophyll cells and by ribose 5-phosphate plus ATP for bundle sheath cells, and the location of phosphoenolpyruvate and ribulose bisphosphate carboxylases in mesophyll and bundle sheath cells, respectively, are in accord with the scheme of C 4 photosynthesis which places the Calvin cycle in the bundle sheath and C 4 acid formation in mesophyll cells. 相似文献
16.
The addition of glyoxylate to tobacco ( Nicotiana tabacum) leaf discs inhibited glycolate synthesis and photorespiration and increased net photosynthetic 14CO 2 fixation. This inhibition of photorespiration was investigated further by studying the effect of glyoxylate on the stimulation of photosynthesis that occurs when the atmospheric O 2 level was decreased from 21 to 3% (the Warburg effect). The Warburg effect is usually ascribed to the increased glycolate synthesis and metabolism that occurs at higher O 2 concentrations. Photosynthesis in control discs increased from 59.1 to 94.7 micromoles of CO 2 per gram fresh weight per hour (a 60% increase) when the O 2 level was lowered from 21 to 3%, while the rate for discs floated on 15 millimolar glyoxylate increased only from 82.0 to 99.7 micromoles of CO 2 per gram fresh weight per hour (a 22% increase). The decrease in the O 2 sensitivity of photosynthesis in the presence of glyoxylate was explained by changes in the rate of glycolate synthesis under the same conditions. The rate of metabolism of the added glyoxylate by tobacco leaf discs was about 1.35 micromoles per gram fresh weight per hour and was not dependent on the O2 concentration in the atmosphere. This rate of metabolism is about 10% the amount of stimulation in the rate of CO2 fixation caused by the glyoxylate treatment on a molar carbon basis. Glyoxylate (10 millimolar) had no effect on the carboxylase/oxygenase activity of isolated ribulose diphosphate carboxylase. Although the biochemical mechanism by which glyoxylate inhibits glycolate synthesis and photorespiration and thereby decreases the Warburg effect is still uncertain, these results show that cellular metabolites can regulate the extent of the Warburg effect. 相似文献
17.
A mass spectrometric method combining 16O/ 18O and 12C/ 13C isotopes was used to quantify the unidirectional fluxes of O 2 and CO 2 during a dark to light transition for guard cell protoplasts and mesophyll cell protoplasts of Commelina communis L. In darkness, O 2 uptake and CO 2 evolution were similar on a protein basis. Under light, guard cell protoplasts evolved O 2 (61 micromoles of O 2 per milligram of chlorophyll per hour) almost at the same rate as mesophyll cell protoplasts (73 micromoles of O 2 per milligram of chlorophyll per hour). However, carbon assimilation was totally different. In contrast with mesophyll cell protoplasts, guard cell protoplasts were able to fix CO 2 in darkness at a rate of 27 micromoles of CO 2 per milligram of chlorophyll per hour, which was increased by 50% in light. At the onset of light, a delay observed for guard cell protoplasts between O 2 evolution and CO 2 fixation and a time lag before the rate of saturation suggested a carbon metabolism based on phospho enolpyruvate carboxylase activity. Under light, CO 2 evolution by guard cell protoplasts was sharply decreased (37%), while O 2 uptake was slowly inhibited (14%). A control of mitochondrial activity by guard cell chloroplasts under light via redox equivalents and ATP transfer in the cytosol is discussed. From this study on protoplasts, we conclude that the energy produced at the chloroplast level under light is not totally used for CO 2 assimilation and may be dissipated for other purposes such as ion uptake. 相似文献
18.
Light-dependent O 2 reduction concomitant with O 2 evolution, ATP formation, and NADP reduction were determined in isolated spinach ( Spinacia oleracea L. var. America) chloroplast lamellae fortified with NADP and ferredoxin. These reactions were investigated in the presence or absence of catalase, providing a tool to estimate the reduction of O 2 to H 2O 2 (Mehler reaction) concomitant with NADP reduction. In the presence of 250 micromolar O 2, O 2 photoreduction, simultaneous with NADP photoreduction, was dependent upon light intensity, ferredoxin, Mn 2+, NADP, and the extent of coupling of phosphorylation to electron flow. In the presence of an uncoupling concentration of NH4+, saturating light intensity (>500 watts/square meter), saturating ferredoxin (10 micromolarity) rate-limiting to saturating NADP (0.2-0.9 millimolarity), and Mn2+ (50-1000 micromolarity), the maxium rates of O2 reduction were 13-25 micromoles/milligram chlorophyll per hour, while concomitant rates of O2 evolution and NADP reduction were 69 to 96 and 134 to 192 micromoles/milligram chlorophyll per hour, respectively. Catalase did not affect the rate of NADPH or ATP formation but decreased the NADPH:O2 ratios from 2.3-2.8 to 1.9-2.1 in the presence of rate-limiting as well as saturating concentrations of NADP. Photosynthetic electron flow at a rate of 31 micromoles O2 evolved/milligram chlorophyll per hour was coupled to the synthesis of 91 micromoles ATP/milligram chlorophyll per hour, while the concomitant rate of O2 reduction was 0.6 micromoles/milligram chlorophyll per hour and was calculated to be associated with an apparent ATP formation of only 2 micromoles/milligram chlorophyll per hour. Thus, electron flow from H2O to O2 did not result in ATP formation significantly above that produced during NADP reduction. 相似文献
19.
Previously, C Baysdorfer and JM Robinson (1985 Plant Physiol 77: 318-320) demonstrated that, in a reconstituted spinach chloroplast system, NADP photoreduction functioning at most maximal rate and reductant demand, was the successful competitor with NO 2− photoreduction for reduced ferredoxin. This resulted in a repression of NO 2− reduction until all NADP available had been almost totally reduced. Further experiments, employing isolated, intact spinach leaf plastids and soybean leaf mesophyll cells, were conducted to examine competition for reductant between CO 2 and NO 2− photoassimilation, in situ. In isolated, intact plastid preparations, regardless of whether the demand for reductant by CO 2 photoassimilation was high (5 millimolar `CO 2') with rates of CO 2 fixation in the range 40 to 90 micromoles CO 2 fixed per hour per milligram chlorophyll, low (0.5 millimolar `CO 2') with rates in the range 5 to 8 micromoles CO 2 per hour per milligram chlorophyll, or zero (no `CO 2'), NO 2− photoreduction displayed equal rates in the range of 8 to 22 micromoles per hour per milligram chlorophyll. In the absence of `CO 2', but in the presence of saturating white light, 3-phosphoglycerate photoreduction at rates of 82 to 127 micromoles per hour per milligram chlorophyll did not repress, and occasionally stimulated concomitant rates of NO 2− reduction which ranged from 23.4 to 38.5. Conversely, in plastid preparations, NO 2− at levels of 50 to 100 micromolar, stimulated plastid CO 2 fixation when `CO 2' was saturating with respect to carboxylation. Further, levels of NO 2− in the range 250 to 2500 micromolar, stimulated soybean leaf mesophyll cell net CO 2 fixation as much as 1.5-fold if `CO 2' was saturating with respect to CO 2 fixation. It appeared likely that, in high light in vivo, CO 2 and NO 2− photoassimilatory processes are not forced to intercompete for reduced ferredoxin in the intact chloroplast. 相似文献
20.
Conditions are described whereby suspensions of Chlorella pyrenoidosa and Netrium digitus photosynthetically biosynthesize and excrete glycolate continuously in high yields. Aminooxyacetic acid, an inhibitor of pyridoxal phosphate-linked enzymes, increased the excretion of glycolate approximately 4-fold in 1 hour (8 millimolar) and 20-fold in 4 hours (40 millimolar) in the presence of 0.2% CO 2 in air. The amount of glycolate excreted in the presence of aminooxyacetate and an atmosphere of 0.2% CO 2 in air equaled or exceeded the amount excreted in 0.2% CO 2 in O 2 minus aminooxyacetate. CO 2 and light were required for glycolate excretion. Aminooxyacetate also stimulated photosynthetic glycolate excretion in an atmosphere of 0.2% CO 2 in nitrogen or helium, although the stimulation was not as great as when air or O 2 was present. The excreted glycolate was converted to H2 and CO2 by the combined action of glycolic oxidase and the formic hydrogenlyase complex found in Escherichia coli in total conversion yields of 80%. 相似文献
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