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1.
Oxygen inhibition of photosynthesis was studied with intact spinach ( Spinacia oleracea L.) chloroplasts which exhibited very high rates of photosynthetic CO 2 reduction and were insensitive to additions of photosynthetic intermediates when CO 2 was available at saturating concentrations. Photosynthetic rates were measured polarographically as O 2 evolution, and the extent of the reduction of substrate was estimated from the amount of O 2 evolved. With CO 2 as substrate, inhibition of photosynthesis by O 2 was dependent on pH. At pH values above 8, rates of O 2 evolution were strongly inhibited by O 2 and only a fraction of the added bicarbonate was reduced before O 2 evolution ceased. The extent of O 2 evolution declined with increasing O 2 concentration and decreasing initial bicarbonate concentration. At pH 7.2, the initial photosynthetic rate was inhibited about 30% at high O 2 levels, but the extent of O 2 evolution was unaffected and most of the added bicarbonate was reduced. Photosynthetic O 2 evolution with 3-phosphoglycerate as substrate was similarly dependent on pH and O 2 concentration. In contrast, there was little effect of O 2 and pH on oxaloacetate-dependent oxygen evolution. Acid-base shift experiments with osmotically shocked chloroplasts showed that ATP formation was not affected by O 2. The results are discussed in terms of a balance between photosynthetic O 2 evolution and O 2 consumption by the ribulose diphosphate oxygenase reaction. 相似文献
4.
Experiments in which [1- 14C]glycolate uptake is carried out in conjunction with measurements of stromal pH indicate that only glycolic acid and not the glycolate anion is crossing the pea ( Pisum sativum var. Progress No. 9, Agway) chloroplast envelope. This mechanism of glycolate transport appears to be too slow to account for observed photorespiratory carbon fluxes in C 3 plants. 相似文献
6.
A sensitive assay based upon fluorescence of scopoletin allowed continuous recording of H 2O 2 production in illuminated intact cells of Anacytis nidulans. Onset of illumination was followed by a 5 to 10 second lag, a burst of very rapid production continuing for up to 5 minutes, and finally a slow and continuing steady rate of H 2O 2 production. Size of the H 2O 2 burst was decreased by 3-(3,4-dichlorophenyl)-1, 1-dimethylurea, by low O 2, and by certain Calvin cycle intermediates; it was increased by high light intensity, CO 2 depletion, Calvin cycle inhibitors (as iodoacetamide), cold shock, carbonyl cyanide m-chlorophenylhydrazone, and certain organic acids as glycolate). The H 2O 2 burst was explained by the following hypothesis: a low potential reductant is produced more rapidly than it can be used in the normal pathway to CO 2 reduction and, instead, reacts with oxygen. H 2O 2 production is regarded as a metabolic defect observable in Anacystis most dramatically during the transition from a very low rate of oxidative dark metabolism to a high rate of photosynthetic metabolism. 相似文献
7.
Intact spinach chloroplasts scavenge hydrogen peroxide witha peroxidase that uses a photoreductant as the electron donor,but the activity of ruptured chloroplasts is very low [Nakanoand Asada (1980) Plant & Cell Physiol. 21 : 1295]. Rupturedspinach chloroplasts recovered their ability to photoreducehydrogen peroxide with the concomitant evolution of oxygen afterthe addition of glutathione and dehydroascorbate (DHA). In rupturedchloroplasts, DHA was photoreduced to ascorbate and oxygen wasevolved in the process in the presence of glutathione. DHA reductase(EC 1.8.5.1
[EC]
) and a peroxidase whose electron donor is specificto L-ascorbate are localized in chloroplast stroma. These observationsconfirm that the electron donor for the scavenging of hydrogenperoxide in chloroplasts is L-ascorbate and that the L-ascorbateis regenerated from DHA by the system: photosystem IferredoxinNADPglutathione.A preliminary characterization of the chloroplast peroxidaseis given. (Received April 16, 1981; Accepted June 3, 1981) 相似文献
8.
Illuminated chloroplasts isolated from SO 2-fumigated spinachleaves accumulated more H 2O 2 than those from non-fumigated ones.This H 2O 2 formation was dependent on light and was inhibitedby DCMU. It also was depressed by cytochrome c and superoxidedismutase (EC 1.15.1.1
[EC]
). The addition of sulfite to rupturedchloroplasts isolated from non-fumigated leaves caused an H 2O 2accumulation that accompanied O 2 uptake. Spinach leaves losttheir catalase (EC 1.11.1.6
[EC]
), ascorbate peroxidase and glutathionereductase (EC 1.6.4.2
[EC]
) activities at the beginning of SO 2 fumigation,when H 2O 2 was accumulated. These results suggest that the accumulationof H 2O 2 in SO 2-fumigated spinach leaves is caused by the increasein O 2production, the precursor for H 2O 2, with a sulfite-mediatedchain reaction at the reducing site of photosystem I, and byinactivation of the H 2O 2 scavenging system. (Received October 7, 1981; Accepted June 16, 1982) 相似文献
9.
A system has been developed for the study of photosynthetic CO 2 fixation by isolated spinach chloroplasts at air levels of CO 2. Rates of CO 2 fixation were typically 20 to 60 micromoles/milligrams chlorophyll per hour. The rate of fixation was linear for 10 minutes but then declined to less than 10% of the initial value by 40 minutes. Ribulose 1,5-bisphosphate (RuBP) levels remained unchanged during this period, indicating that they were not the cause for the decline. The initial activity of the RuBP carboxylase in the chloroplast was high for 8 to 10 minutes and then declined similar to the rate of CO 2 fixation, suggesting that the decline in CO 2 fixation may have been caused by deactivation of the enzyme. 相似文献
10.
Conditions for optimal CO 2 fixation and malate decarboxylation by isolated bundle sheath chloroplasts from Zea mays were examined. The relative rates of these processes varied according to the photosynthetic carbon reduction cycle intermediate provided. Highest rates of malate decarboxylation, measured as pyruvate formation, were seen in the presence of 3-phosphoglycerate, while carbon fixation was highest in the presence of dihydroxyacetone phosphate; only low rates were measured with added ribose-5-phosphate. Chloroplasts exhibited a distinct phosphate requirement and this was optimal at a level of 2 millimolar inorganic phosphate in the presence of 2.5 millimolar 3-phosphoglycerate, dihydroxyacetone phosphate, or ribose-5-phosphate. Malate decarboxylation and CO 2 fixation were stimulated by additions of AMP, ADP, or ATP with half-maximal stimulation occurring at external adenylate concentrations of about 0.15 millimolar. High concentrations (>1 millimolar) of AMP were inhibitory. Aspartate included in the incubation medium stimulated malate decarboxylation and CO 2 assimilation. In the presence of aspartate, the apparent Michaelis constant (malate) for malate decarboxylation to pyruvate by chloroplasts decreased from 6 to 0.67 millimolar while the calculated Vmax for this process increased from 1.3 to 3.3 micromoles per milligram chlorophyll. Aspartate itself was not metabolized. It was concluded that the processes mediating the transport of phosphate, 3-phosphoglycerate, and dihydroxyacetone phosphate transport on the one hand, and also of malate might differ from those previously described for chloroplasts from C 3 plants. 相似文献
11.
Illuminated intact spinach chloroplasts decomposed one moleculeof H 218O 2 which resulted in the evolution of a half moleculeof 16O 2, but little 18O 2. The chloroplasts showed the same rateof photoreduction of 18C 2 as that of the evolution of 16O 2 withoutaccumulation of H 218O 2. These reactions were suppressed by DCMU,and also by several inhibitors of ascorbate peroxidase and dehydroascorbateand monodehydroascorbate reductases in chloroplasts. These observationsindicate that the hydrogen peroxide produced in chloroplastsis reduced to water by a peroxidase using a photoreductant asthe electron donor. The hydrogen peroxide scavenging systemof chloroplasts was inactivated if hydrogen peroxide was addedin the dark, but not if added during the light. (Received May 4, 1984; Accepted July 10, 1984) 相似文献
12.
Intact spinach ( Spinacia oleracea L.) chloroplasts, when pre-illuminated at 4 millimoles quanta per square meter per second for 8 minutes in a CO 2-free buffer at 21% O 2, showed a decrease (30-70%) in CO 2-dependent O 2 evolution and 14CO 2 uptake. This photoinhibition was observed only when the O 2 concentration and the quantum fluence rate were higher than 4% and 1 millimole per square meter per second, respectively. There was only a small decrease in the extent of photoinhibition when the CO 2 concentration was increased from 0 to 25 micromolar during the treatment, but photoinhibition was abolished when the CO 2 concentration was increased to 30 micromolar. Addition of small quantities of P-glycerate (40-200 micromolar) or glycerate (160 micromolar) was found to prevent photoinhibition. Other intermediates of the Calvin cycle (fructose-6-P, fructose-1,6-P, ribose-5-P, ribulose-5-P) also prevented photoinhibition to various extents. Oxaloacetate was not effective in preventing photoinhibition in these chloroplasts. The amount of O 2 evolved during treatments with 3-P-glycerate or glycerate was no more than 65% of that measured in the presence of low CO 2 concentrations (9-12 micromolar) which did not prevent photoinhibition. In all cases, the extent to which photoinhibition was prevented by these metabolites was not correlated to the amount of O 2 evolved during the photoinhibitory treatment. It is concluded that in these chloroplasts the prevention of the O 2-dependent photoinhibition of light saturated CO 2 fixation capacity is not linked to the dissipation of excitation energy via the photosynthetic electron transport nor to ATP utilization. The requirement of O 2 for photoinhibition of CO 2 fixation capacity in isolated chloroplasts may be explained by an effect of O 2 in allowing metabolic depletion of Calvin cycle intermediates. 相似文献
13.
Photoassimilation of 14CO 2 by intact chloroplasts from the Crassulacean acid metabolism plant Sedum praealtum was investigated. The main water-soluble, photosynthetic products were dihydroxyacetone phosphate (DHAP), glycerate 3-phosphate (PGA), and a neutral saccharide fraction. Only a minor amount of glycolate was produced. A portion of neutral saccharide synthesis was shown to result from extrachloroplastic contamination, and the nature of this contamination was investigated with light and electron microscopy. The amount of photoassimilated carbon partitioned into starch increased at both very low and high concentrations of orthophosphate. High concentrations of exogenous PGA also stimulated starch synthesis. DHAP and PGA were the preferred forms of carbon exported to the medium, although indirect evidence suported hexose monophosphate export. The export of PGA and DHAP to the medium was stimulated by high exogenous orthophosphate, but depletion of chloroplastic reductive pentose phosphate intermediates did not occur. As a result only a relatively small inhibition in the rate of CO2 assimilation occurred. The rate of photoassimilation was stimulated by exogenous PGA, ribose 5-phosphate, fructose 1,6-bisphosphate, fructose 6-phosphate, and glucose 6-phosphate. Inhibition occurred with phosphoenolpyruvate and high concentrations of PGA and ribose 5-phosphate. PGA inhibition did not result from depletion of chloroplastic orthophosphate or from inhibition of ribulose 1,5-bisphosphate carboxylase. Exogenous PGA and phosphoenolpyruvate were shown to interact with the orthophosphate translocator. 相似文献
14.
Uptake of l-[1- 14C]ascorbate by intact ascorbate-free spinach ( Spinacia oleracea L. cv Vital r) chloroplasts has been investigated using the technique of silicone oil filtering. Rates greater than 100 micromoles per milligram chlorophyll per hour (external concentration, 10 millimolar) of ascorbate transport were observed. Ascorbate uptake into the sorbitol-impermeable space (stroma) followed the Michaelis-Menten-type characteristic for substrate saturation. A Km of 18 to 40 millimolar was determined. Transport of ascorbate across the chloroplast envelope resulted in an equilibrium of the ascorbate concentrations between stroma and medium. A pH optimum of 7.0 to 7.5 and the lack of alkalization of the medium upon ascorbate uptake suggest that only the monovalent ascorbate anion is able to cross the chloroplast envelope. The activation energy of ascorbate uptake was determined to be 65.8 kilojoules (16 kilocalories) per mole (8 to 20°C). Interference of ascorbate transport with substrates of the phosphate or dicarboxylate translocator could not be detected, but didehydroascorbate was a competitive inhibitor. Preloading of chloroplasts with didehydroascorbate resulted in an increase of Vmax but did not change the Km for ascorbate. Millimolar concentrations of the sulfhydryl reagent p-chloromercuriphenyl sulfonate inhibited ascorbate uptake. The data are interpreted in terms of ascorbate uptake into chloroplasts by the mechanism of facilitated diffusion mediated by a specific translocator. 相似文献
15.
Alkali-salinity exerts severe osmotic, ionic, and high-pH stresses to plants. To understand the alkali-salinity responsive mechanisms underlying photosynthetic modulation and reactive oxygen species (ROS) homeostasis, physiological and diverse quantitative proteomics analyses of alkaligrass ( Puccinellia tenuiflora) under Na2CO3 stress were conducted. In addition, Western blot, real-time PCR, and transgenic techniques were applied to validate the proteomic results and test the functions of the Na 2CO 3-responsive proteins. A total of 104 and 102 Na 2CO 3-responsive proteins were identified in leaves and chloroplasts, respectively. In addition, 84 Na 2CO 3-responsive phosphoproteins were identified, including 56 new phosphorylation sites in 56 phosphoproteins from chloroplasts, which are crucial for the regulation of photosynthesis, ion transport, signal transduction, and energy homeostasis. A full-length PtFBA encoding an alkaligrass chloroplastic fructose-bisphosphate aldolase (FBA) was overexpressed in wild-type cells of cyanobacterium Synechocystis sp. Strain PCC 6803, leading to enhanced Na 2CO 3 tolerance. All these results indicate that thermal dissipation, state transition, cyclic electron transport, photorespiration, repair of photosystem (PS) II, PSI activity, and ROS homeostasis were altered in response to Na 2CO 3 stress, which help to improve our understanding of the Na 2CO 3-responsive mechanisms in halophytes. 相似文献
17.
The feasibility of a luminol-dependent chemiluminescence analysis of hydrogen peroxide production by intact wheat seedlings using a KhL-003 chemiluminometer was determined. It was shown that the minimal H 2O 2 concentration that can be detected in a 0.5-ml sample with this instrument is 0.125 µM. Analysis of biological activity of a mixture of chitooligosaccharides with molecular masses from 5 to 10 kD and acetylation degree of 65% demonstrated that, at a concentration of 1 µg/ml, they induce rapid overproduction of H 2O 2 in roots of 3-day-old wheat seedlings. 相似文献
18.
Illuminated suspensions of Anacystis nidulans, supplied with saturating concentrations of CO 2 evolved O 2 at a greater rate when nitrate was simultaneously present. The extent of the stimulation of noncyclic electron flow induced by nitrate was dependent on light intensity, being maximal under light saturating conditions. Accordingly, nitrate depressed the rate of CO 2 fixation at limiting but not at saturating light, this depression reflecting the competition between both processes for assimilatory power. In contrast, ammonium stimulated CO 2 fixation at any light intensity assayed, the stimulation being dependent on the incorporation of ammonium to carbon skeletons. The positive effect of ammonium on CO 2 fixation also appeared to occur when nitrate was the nitrogen source, since with either nitrogen source an increase in the incorporation of newly fixed carbon into acid-soluble metabolites took place. From these results, the in vivo partitioning of assimilatory power between photosynthetic nitrogen and carbon assimilation and the quantitative and qualitative effects of inorganic nitrogen assimilation on CO 2 fixation are discussed. 相似文献
19.
A method for purification and crystallization, and some properties of ribonuclease from Aspergillus sp. [EC 2.7.7.17] (RNase L) are reported. The purification procedure consisted of six steps, including acetone precipitation, column chromatographies on Duolite A–2 and DEAE-cellulose, repeated chromatography on DEAE-Sephadex A–50 column and affinity chromatography on 5’-AMP-Sepharose 4B column. Crystallization was performed by the dialysis against ammonium sulfate solution at 60% saturation.The crystalline enzyme was shown to be homogeneous by polyacrylamide disc electrophoresis and ultracentrifugation. Svedberg value of the crystalline enzyme was 4.2. The enzyme was the most active at pH 3.5 and 60~65°C, and it was inhibited markedly with Fe. 3+RNase L has no absolute base specificity, and produces four kinds of 3’-mononucleotides from yeast RNA. However, the susceptibility of four nucleotide residues to RNase L increases in the order; G < A < C < U and is quite different from those of RNase T 2, RNase M and RNase R. 相似文献
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