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1.
Abstract A simple mechanical method for the rapid isolation of chloroplasts with high rates of photosynthesis from young leaves of oat ( Avena sativa L.) was described. The photosynthetic activity of these chloroplasts was stable for at least 2 h with rates of CO 2-dependent O 2 evolution of 30–40 μmol g 1 Chl s 1. The photosynthetic properties of these chloroplasts were similar to those reported for spinach and pea chloroplasts isolated by mechanical disruption. The pH optimum for photosynthetic O 2 evolution was pH 7.6. The induction time was 0.5–2 min. Maximal rates of photosynthetic O 2 evolution in these chloroplast preparations were obtained in the absence of both divalent cations and EDTA. Addition of divilent cations strongly inhibited photosynthesis which could be partially restored by the subsequent addition of EDTA. But when these cations were not present in the assay medium the addition of EDTA greater than 1 mol m 3 decreased photosynthetic activity. The optimal orthophosphate concentration required for photosynthesis in these chloroplast preparations was 0.2–0.3 mol m 3. In contrast, the addition of pyrophosphate either in the light or dark inhibited photosynthesis. In a comparative study, chloroplasts were also isolated from oat and wheat ( Triticum aestivum L., cultivar Hybrid C306) protoplasts. These chloroplast preparations were found to have properties similar to those determined for oat chloroplasts isolated by the mechanical method reported above. 相似文献
2.
Chloroplasts, isolated from protoplasts of the green alga, Chlorella ellipsoidea, were estimated to be 99% intact by the ferricyanide-reduction assay, and gave CO 2 and PGA-dependent rates of O 2 evolution of 64.5 to 150 micromoles per milligram of chlorophyll per hour, that is 30 to 70% of the photosynthetic activity of the parent cells. Intact chloroplasts showed no carbonic anhydrase activity, but it was detected in preparations of ruptured organelles. Rates of photosynthesis, measured in a closed system at pH 7.5, were twice the calculated rate of CO 2 supply from the uncatalyzed dehydration of HCO 3− indicating a direct uptake of bicarbonate by the intact chloroplasts. Mass spectrometric measurements of CO 2 depletion from the medium on the illumination of chloroplasts indicate the lack of an active CO 2 transport across the chloroplast envelope. 相似文献
3.
Various physiological characteristics of photosynthesis in the unicellular red alga Porphyridium cruentum Naegeli have been investigated. The rate of photosynthesis was optimal at 25° C and pH 7.5 and was not inhibited by 21% oxygen over a temperature range of 5 to 35° C. Kinetics of whole cell photosynthesis as a function of substrate concentration gave a K 1/2, (CO 2) of 0.3 μM. CO 2 compensation point, measured in a closed system at pH 7.5, was a constant 6.7 m?L · L ?1 over the temperature range 15 to 30° C and was unaffected by O 2 concentration. Whole cell photosynthesis, measured in a closed system at alkaline pH, showed that the rates of oxygen evolution were greatly in excess of the rate of CO 2 supply from the spontaneous dehydration of HCO 3? in the medium. This indicates that bicarbonate is utilized by the cell to support this photosynthetic rate. These physiological characteristics of Porphyridium cruentum are consistent with the hypothesis that this alga transports bicarbonate across the plasmalemma. 相似文献
4.
This study examines the effect of antimycin A and nitrite on 14CO 2 fixation in intact chloroplasts isolated from spinach ( Spinacia oleracea L.) leaves. Antimycin A (2 micromolar) strongly inhibited CO 2 fixation but did not appear to inhibit or uncouple linear electron transport in intact chloroplasts. The addition of small quantities (40-100 micromolar) of nitrite or oxaloacetate, but not NH 4Cl, in the presence of antimycin A restored photosynthesis. Antimycin A inhibition, and the subsequent restoration of photosynthetic activities by nitrite or oxaloacetate, was observed over a wide range of CO 2 concentration, light intensity, and temperature. High O 2 concentration (up to 240 micromolar) did not appear to influence the extent of the inhibition by antimycin A, nor the subsequent restoration of photosynthetic activity by nitrite or oxaloacetate. Studies of O 2 exchanges during photosynthesis in cells and chloroplasts indicated that 2 micromolar antimycin A stimulated O 2 uptake by about 25% while net O 2 evolution was inhibited by 76%. O 2 uptake in chloroplasts in the presence of 2 micromolar antimycin A was 67% of total O 2 evolution. These results suggest that only a small proportion of the O 2 uptake measured was directly linked to ATP generation. The above evidence indicates that cyclic photophosphorylation is the predominant energy-balancing reaction during photosynthesis in intact chloroplasts. On the other hand, pseudocyclic O 2 uptake appears to play only a minimal role. 相似文献
5.
The catalase activity of unwashed preparations containing intact spinach ( Spinacia oleracea L.) chloroplasts is inhibited both by cyanide and by azide at concentrations which also cause inhibition of photosynthetic CO 2- dependent O 2 evolution. Aminotriazole can also be used to inhibit this contaminant catalase, and in this case inhibition of catalase can be achieved at aminotriazole concentrations which have little effect on the rate of photosynthetic CO2 fixation. Aminotriazole may be used as a specific inhibitor of catalase in order to demonstrate inhibition of photosynthesis by added H2O2. It is therefore concluded that inhibition of photosynthesis by cyanide and azide does not necessarily result from inhibition of catalase in the chloroplast preparation, and that intact chloroplasts do not produce inhibitory concentrations of H2O2 under the best experimental conditions for CO2 fixation. 相似文献
6.
Cyanide and azide inhibit photosynthesis and catalase activity of isolated, intact spinach ( Spinacia oleracea) chloroplasts. When chloroplasts are illuminated in the presence of CN − or N 3−, accumulation of H 2O 2 is observed, parallel to inhibition of photosynthesis. Photosynthetic O 2 evolution is inhibited to the same extent, under saturating light, whether CO 2 or phosphoglycerate is present as electron acceptor. 相似文献
7.
The ability of the seagrass Zostera muelleri Irmisch ex Aschers. to use HCO 3− as well as CO 2 for photosynthesis was investigated by measuring photosynthetic O 2 evolution over a range of pH values. It was found that the apparent Km CO 2 fell from 0.128 mM at pH 7.9 to 0.016 mM at pH 9.1 indicating that HCO 3− as well as CO 2 may act as a substrate for photosynthesis.The true Km CO 2 could not be determined due to inhibition of photosynthesis at pHs less than 7.8 Km CO 2 must be at least 0.128 mM, the apparent Km at pH 7.9, and is probably of the order of 0.200 mM CO 2, the same as that reported for other marine plants. Km HCO 3−1 is about 20 mM when CO 2-dependent photosynthesis is minimal. Such a high Km HCO 3− resembles values reported for freshwater, rather than marine plants.Photosynthetic O 2 evolution is not saturated with respect to total inorganic carbon in natural seawater (pH 8.2). It is suggested that the distinctive shoulder from pH 8.1 to 8.5 in the pH profile of photosynthetic O 2 evolution at a constant concentration of inorganic carbon is caused by an effect of pH on HCO 3− uptake. The effect of pH on HCO 3− uptake was determined by constructing a pH profile of photosynthesis at constant HCO 3− concentration, and subtracting the estimated contribution of CO 2 to photosynthesis from this rate. The resultant curve has a maximum at pH 8.4 and declines sharply at pHs less than 8. 相似文献
8.
Magnesium was most inhibitory to photosynthetic reactions by intact chloroplasts when the magnesium was added in the dark before illumination. Two millimolar MgCl 2, added in the dark, inhibited CO 2-dependent O 2 evolution by Hordeum vulgare L. and Spinacia oleracea L. (C 3 plants) chloroplasts 70 to 100% and inhibited (pyruvate + oxaloacetate)-dependent O 2 evolution by Digitaria sanguinalis L. (C 4 plant) mesophyll chloroplasts from 80 to 100%. When Mg 2+ was added in the light, O 2 evolution was reduced only slightly. O 2 evolution in the presence of phosphoglycerate was less sensitive to Mg 2+ inhibition than was CO 2-dependent O 2 evolution. Magnesium prevented the light activation of several photosynthetic enzymes. Two millimolar Mg2+ blocked the light activation of NADP-malate dehydrogenase in D. sanguinalis mesophyll chloroplasts, and the light activation of phosphoribulokinase, NADP-linked glyceraldehyde-3-phosphate dehydrogenase, and fructose 1,6-diphosphatase in barley chloroplasts. The results suggest that Mg2+ inhibits chloroplast photosynthesis by preventing the light activation of certain enzymes. 相似文献
9.
Intact chloroplasts of wheat ( Triticum aestivum) were isolated from mesophyll protoplasts. With decreasing concentrations of bicarbonate from 10 to 0.3 millimolar (pH 8.0), the optimal concentration of orthophosphate (Pi) for photosynthetic O 2 evolution decreased from a value of 0.1 to 0.2 millimolar to 0 to 0.025 millimolar. The extremely low Pi optimum for photosynthesis at the low bicarbonate levels of 0.3 millimolar was increased by lowering the O 2 concentration from 253 (21% gas phase) to 72 micromolar (6% gas phase). The relative amount of glycolate and dihydroxyacetone phosphate (DHAP) synthesized under high and low levels of bicarbonate and varying levels of Pi was determined. At low levels of bicarbonate, glycolate was the main product, whereas at high bicarbonate levels, DHAP was the main product. Most of the DHAP and glycolate was found in the extrachloroplastic fraction. 相似文献
10.
A new approach to measurements of photosynthesis by isolated chloroplasts has been devised. Intact isolated chloroplasts were trapped in the cavities of membrane filters. The thin layers of chloroplasts so obtained were assayed for O 2 evolution and CO 2 assimilation in leaf-chambers. Photosynthetic gas exchange could be demonstrated to take place either in a closed or a flow-through system. The chloroplasts were morphologically intact as shown by light or scanning electron microscopy and displayed stable rates of photosynthesis in the presence of phosphate and alkaline phosphatase. The methods described open the way to in vitro measurement of photosynthesis, by chloroplasts under conditions more closely resembling those in leaves. 相似文献
11.
In order to study the relative contributions of the autocatalytic increase in the level of substrates and the light activation of enzymes to the control of the induction phase or “lag” in wheat chloroplasts, we measured the light-induced reductive activation of fructose 1,6-bisphosphatase, phosphoglycerate kinase, NADP +-dependent glyceraldehyde-phosphate dehydrogenase, ribulose 1,5-bisphosphate carboxylase, and phosphoribulokinase in isolated chloroplasts. Each was rapidly activated to levels more than adequate to support the maximum rate of photosynthesis. Induction in wheat chloroplasts is characterized by a period of about 1 min during which no O 2 is evolved. If small quantities of intermediates such as dihydroxyacetone phosphate (DHAP) or 3-phosphoglycerate (PGA) are added, maximum rates of photosynthesis are achieved within the first minute of illumination. The presence of PGA did not affect the activation of any of the above-mentioned enzymes. Each of the enzymes was therefore capable of sustaining maximum rates of photosynthesis in the presence of PGA, even though there was no O 2 evolution from those chloroplasts incubated with CO 2 alone as substrate. The inclusion of PGA did not give rise to abnormally high levels of DHAP, FBP, or fructose 6-phosphate in the stroma. We conclude that the levels of substrates or cofactors are the principal, if not the sole, determinants of the rate of photosynthetic carbon assimilation during induction in wheat chloroplasts. 相似文献
12.
Photosynthetic carbon uptake of the tropical seagrass Thalassia hemprichii (Ehrenb.) Aschers was studied by several methods. Photosynthesis in buffered seawater in media in the range of pH 6 to pH 9 showed an exponentially increasing rate with decreasing pH, thus indicating that free CO 2 was a photosynthetic substrate. However, these experiments were unable to determine whether photosynthesis at alkaline pH also contained some component due to HCO 3− uptake. This aspect was further investigated by studying photosynthetic rates in a number of media of varying pH (7.8-8.61) and total inorganic carbon (0.75-13.17 millimolar). In these media, photosynthetic rate was correlated with free CO 2 concentration and was independent of the HCO 3− concentration in the medium. Short time-course experiments were conducted during equilibration of free CO 2 and HCO 3− after injection of 14C labeled solution at acid or alkaline pH. High initial photosynthetic rates were observed when acidic solutions (largely free CO 2) were used but not with alkaline solutions. The concentration of free CO 2 was found to be a limiting factor for photosynthesis in this plant. 相似文献
13.
A sterile continuous turbidostat culture in a 2-1 fermenter was used to systematically measure the gas exchange rates of Anacystis nidulans in a highly turbulent system under strictly controlled environmental conditions. An extensive physiological characterization of Anacystis is given in terms of photosynthesis rates (CO 2 uptake and O 2 evolution) and dark respiration rates as function of different parameters such as stirrer speed, temperature, CO 2 and O 2 concentration, light intensity, culture density and pH. Steady state ATP levels and apparent photophosphorylation rates complete the performance data. The dependence of the photosynthetic quotient from the parameters enables a physiological characterization of the light dependent nitrate assimilation. 相似文献
14.
In isolated intact chloroplasts, maximal rates of photosynthetic O 2 evolution (in saturating HCO ?3) are associated with a critical transthylakoid proton gradient as a result of the stoichiometric consumption of 2 mol NADPH and 3 mol ATP/mol CO 2 fixed. Studies with the fluorescent probe 9-aminoacridine reveal that in the illuminated steady state the critical ΔpH is 3.9.CO 2-dependent O 2 evolution is inhibited by increases of 0.1–0.2 in ΔpH that occur when catalase is omitted from the medium, NO ?2 is included as an electron acceptor, or when chloroplasts are illuminated under low partial pressures of O 2. Low concentrations of antimycin (0.33 μM) or NH 4Cl (0.33 mM) decrease ΔpH and relieve this inhibition of electron flow. The energy transfer inhibitor quercetin lowers the high ATP/ADP ratio associated with these conditions, but does not lower ΔpH or relieve the inhibition.A decrease of ΔpH below 3.9 by weaker illumination, millimolar levels of NH 4Cl or micromolar levels of antimycin, results in lower rates of photosynthesis owing to limitation by the phosphorylation rate.These findings show that in absence of rate limitation by the carbon cycle, the extent of thylakoid energization is related to the ratio of ATP to NADPH production and in turn, the rate of CO 2 assimilation. 相似文献
15.
Two strains of marine Synechococcus possessed a much greater potential for photorespiration than other marine algae we have studied. This conclusion was based
on the following physiological and biochemical characteristics: a) a light-dependent O 2 inhibition of photosynthetic CO 2 assimilation at atmospheric O 2 concentrations. The degree of inhibition was dependent on the relative concentrations of dissolved O 2 and CO 2, being greatest at 100% O 2 with no extra bicarbonate added to the medium; b) actively photosynthesizing cells had high levels of ribulose-1,5-bisphosphate
carboxylase compared with phosphoenolpyruvate carboxylase; ribulose-1,5-bisphosphate oxygenase activities were three times
greater than ribulose-1,5-bisphosphate carboxylase activities; c) cells photosynthesizing in 21% O 2, showed significant 14C-labelling of phosphoglycolate and glycolate and the percentage of total carbon-14 incorporated into these two compounds
increased when the O 2 concentration was 100%; d) at 100% O 2, there was a post-illumination enhanced rate of O 2 consumption, which was three times greater than dark respiration, and the rate declined with increasing bicarbonate concentrations.
The inhibitory effect of O 2 on photosynthesis did not appear to be solely due to photorespiration, since O 2 inhibition of photosynthetic O 2 evolution was much greater than that of photosynthetic CO 2 assimilation. Also, O 2 inhibition of photosynthetic O 2 evolution declined only slightly with decreasing light intensities, while the inhibition of CO 2 assimilation declined rapidly with decreasing light intensity. 相似文献
16.
Abstract A comparison of some of the methods used to determine whether aquatic plants have the ability to utilize bicarbonate ions as a source of inorganic carbon for photosynthesis has been applied to the intertidal macroalga Ascophyllum nodosum. These include: observing photosynthesis at a high pH (below the alga's CO 2 compensation point), pH compensation point determinations, comparing the photosynthetic characteristics at low pH (5.20) and at high pH (7.95), estimating the maximal rates at which CO 2 can diffuse through the unstirred layer and the rate at which CO 2 can be produced from bicarbonate dehydration in the unstirred layer. All indicated that Ascophyllum nodosum can use bicarbonate ions for photosynthesis, though some were not always consistent. Calculating the total inorganic carbon concentration from pH measurements and acidification CO 2 determinations revealed that the assumption that the alkalinity remains constant during pH drift experiments is not always valid. 相似文献
17.
Chloroplast from greening potato tuber showed good photosynthetic capacity. The evolution of O 2 was dependent upon the intensity of light. A light intensity of 30 lux gave maximum O 2 evolution. At higher intensities inhibition was observed. The presence of bicarbonate in the reaction mixture was essential for O 2 evolution. NADP was found to be a potent inhibitor of O 2 evolution in this system. NADP and 3-(3,4-dichlorophenyl)-1,1-dimethyl urea (DCMU) inhibited the O 2 evolution completely at a 3 μm concentration level, which was reversed by oxidized 2,6-dichlorophenol-indophenol (DCIP). Cyanide (CN)-treated chloroplasts showed full O 2 evolution capacity, when a lipophilic electron acceptor like N-tetramethyl- p-phenylenediamine (TMPD) or DCIP was used along with ferricyanide. Ferricyanide alone showed only 20% reduction. NADP or DCMU could inhibit O 2 evolution only when TMPD was the acceptor but not with DCIP. Photosystem II (PS II) isolated from these chloroplasts also showed inhibition by NADP or DCMU and its reversal by DCIP. Here also the evolution of O 2 with only TMPD as acceptor was sensitive to NADP or DCMU. In the presence of added silicotungstate in PS II NADP or DCMU did not affect ferricyanide reduction or oxygen evolution. The chloroplasts were able to bind exogenously added NADP to the extent of 120 nmol/mg chlorophyll. It is concluded that the site of inhibition of NADP is the same as in DCMU, and it is between the DCIP and TMPD acceptor site in the electron transport from the quencher (Q) to plastoquinone (PQ). 相似文献
18.
Inorganic carbon acquisition has been investigated in the marine haptophyte Isochrysis galbana. External carbonic anhydrase (CA) was present in air‐grown (0.034% CO 2) cells but completely repressed in high (3%) CO 2‐grown cells. External CA was not inhibited by 1.0 mM acetazolamide. The capacity of cells to take up bicarbonate was examined by comparing the rate of photosynthetic O 2 evolution with the calculated rate of spontaneous CO 2 supply; at pH 8.2 the rates of O 2 evolution exceeded the CO 2 supply rate 14‐fold, indicating that this alga was able to take up HCO 3 ? . Monitoring CO 2 concentrations by mass spectrometry showed that suspensions of high CO 2‐grown cells caused a rapid drop in the extracellular CO 2 in the light and addition of bovine CA raised the CO 2 concentration by restoring the HCO 3 ? ‐CO 2 equilibrium, indicating that cells were maintaining the CO 2 in the medium below its equilibrium value during photosynthesis. A rapid increase in extracellular CO 2 concentration occurred on darkening the cells, indicating that the cells had accumulated an internal pool of unfixed inorganic carbon. Active CO 2 uptake was blocked by the photosynthetic electron transport inhibitor 3‐(3′,4′‐dichlorphenyl)‐1,1‐dimethylurea, indicating that CO 2 transport was supported by photosynthetic reactions. These results demonstrate that this species has the capacity to take up HCO 3 ? and CO 2 actively as sources of substrate for photosynthesis and that inorganic carbon transport is not repressed by growth on high CO 2, although external CA expression is regulated by CO 2 concentration. 相似文献
19.
Cotton ( Gossypium hirsutum L. cv Stoneville 213) was grown at 350 and 1000 microliters per liter CO 2. The plants grown at elevated CO 2 concentrations contained large starch pools and showed initial symptoms of visible physical damage. Photosynthetic rates were lower than expected based on instantaneous exposure to high CO 2. A group of plants grown at 1000 microliters per liter CO2 was switched to 350 microliters per liter CO2. Starch pools and photosynthetic rates were monitored in the switched plants and in the two unswitched control groups. Photosynthetic rates per unit leaf area recovered to the level of the 350 microliters per liter CO2 grown control group within four to five days. To assess only nonstomatal limitations to photosynthesis, a measure of photosynthetic efficiencies was calculated (moles CO2 fixed per square meter per second per mole intercellular CO2). Photosynthetic efficiency also recovered to the levels of the 350 microliters per liter CO2 grown controls within three to four days. Recovery was correlated to a rapid depletion of the starch pool, indicating that the inhibition of photosynthesis is primarily a result of feedback inhibition. However, complete recovery may involve the repair of damage to the chloroplasts caused by excessive starch accumulation. The rapid and complete reversal of photosynthetic inhibition suggests that the appearance of large, strong sinks at certain developmental stages could result in reduction of the large starch accumulations and that photosynthetic rates could recover to near the theoretical capacity during periods of high photosynthate demand. 相似文献
20.
Photosynthetic (oxygen evolution) and growth (biomass increase) responses to ambient pH and inorganic carbon (Ci) supply were
determined for Porphyralinearis grown in 0.5 L glass cylinders in the laboratory, or in 40 L fibreglass outdoor tanks with running seawater. While net photosynthetic
rates were uniform at pH 6.0–8.0, dropping only at pH 8.7, growth rates were significantly affected by pH levels other than
that of seawater (c. pH 8.3). In glass cylinders, weekly growth rates averaged 76% at external pH 8.0, 13% at pH 8.7 and 26%
at pH 7.0. Photosynthetic O 2 evolution on a daily basis(i.e. total O 2 evolved during day time less total O 2 consumed during night time) was similar to the growth responses at all experimental pH levels, apparently due to high dark
respiration rates measured at acidic pH. Weekly growth rates averaged 53% in algae grown in fibreglass tanks aerated with
regular air (360 mg L -1 CO 2) and 28% in algae grown in tanks aerated with CO 2-enriched air (750 mg L -1 CO 2). The pH of the seawater medium in which P. linear is was grown increased slightly during the day and only rarely reached 9.0. The pH at the boundary layer of algae submerged
in seawater increased in response to light reaching, about pH 8.9 within minutes, or remained unchanged for algae submerged
in a CO 2-free artificial sea water medium. Photosynthesis of P. linearissaturated at Ci concentrations of seawater (K 0.5560 μM at pH 8.2) and showed low photosynthetic affinity for CO 2(K 0.5 61 μM) at pH 6.0. It is therefore concluded that P. linearisuses primarily CO 2 with HCO 3
- being an alternative source of Ci for photosynthesis. Its fast growth could be related to the enzyme carbonic anhydrase whose
activity was detected intra- and extracellularly.
This revised version was published online in August 2006 with corrections to the Cover Date. 相似文献
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