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1.
The inhibition of photosynthesis after supplying glucose to detached leaves of spinach (Spinacia oleracea L.) was used as a model system to search for mechanisms which potentially contribute to the sink regulation of photosynthesis. Detached leaves were supplied with 50 mM glucose or water for 7 d through the transpiration stream, holding the leaves in low irradiance (16 mol photons · m–2 · s–1) and a cycle of 9 h light/15 h darkness to prevent any endogenous accumulation of carbohydrate. Leaves supplied with water only showed marginal changes of photosynthesis, respiration, enzyme levels or metabolites. When leaves were supplied with 50 mM glucose, photosynthesis was gradually inhibited over several days. The inhibition was most marked when photosynthesis was measured in saturating irradiance and ambient CO2, less marked in saturating irradiance and saturating CO2, and least marked in limiting irradiance. There was a gradual loss of ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco) protein, fructose-1,6-bisphosphatase, NADP-glyceraldehyde-3-phosphate dehydrogenase and chlorophyll. The inhibition of photosynthesis was accompanied by a large decrease of glycerate-3-phosphate, an increase of triose-phosphates and fructose-1,6-bisphospate, and a small decrease of ribulose-1,5-bisphosphate. The stromal NADPH/NADP ratio increased (as indicated by increased activation of NADP-malate dehydrogenase), and the ATP/ADP ratio increased. Chlorophyll-fluorescence analysis indicated that thylakoid energisation was increased, and that the acceptor side of photosystem II was more reduced. Similar results were obtained when glucose was supplied by floating leaf discs in low irradiance on glucose solution, and when detached spinach leaves were held in high light to produce an endogenous accumulation of carbohydrate. Feeding glucose also led to an increased rate of respiration. This was not accompanied by any changes of pyruvate kinase, phosphofructokinase, or pyrophosphate: fructose-6-phosphate phosphotransferase activity. There was a decrease of phosphoenolpyruvate, glycerate-3-phosphate and glycerate-2-phosphate, an increase of pyruvate and triose-phosphates, and an increased ATP/ADP ratio. These results show (i) that accumulation of carbohydrate can inhibit photosynthesis via a long-term mechanism involving a decrease of Rubisco and other Calvin-cycle enzymes and (ii) that respiration is stimulated due to an unknown mechanism, which increases the utilisation of phosphoenolpyruvate.Abbreviations and Symbols Ci
CO2 concentration in the air space within the leaf
- Fm
fluorescence yield with a saturating pulse in dark-adapted material
- Fo
ground level of fluorescence using a weak non-actinic modulated beam in the dark
- Fru1,6bisP
fructose-1,6-bisphosphate
- Fru1,6Pase
fructose-1,6-bisphosphatase
- Fru2,6bisP
fructose-2,6-bisphosphate
- IRGA
infrared gas analyser
- NAD-MDH
NAD-dependent malate dehydrogenase
- NADP-MDH
NADP-dependent malate dehydrogenase
- NADP-GAPDH
NADP-dependent glyceraldehyde-3-phosphate dehydrogenase
- PEP
phosphoenolpyruvate
- PFK
phospho-fructokinase
- PFP
pyrophospate: fructose-6-phosphate-phosphotransferase
- 3-PGA
glycerate-3-phospate
- Pi
inorganic phosphate
- Ru1,5bisP
ribulose 1,5-bisphosphate
- Rubisco
ribulose-1,5-bisphosphate carboxylase-oxygenase
- triose-phosphates
sum of glyceraldehyde-3-phosphate and dihydroxyacetone phosphate
This research was supported by the Deutsche Forschungsgemeinschaft (SFB 137). 相似文献
2.
The objective of this study was to determine whether exposure of plants to ozone (O3) increased the foliar levels of glucose, glucose sources, e.g., sucrose and starch, and glucose-6-phosphate (G6P), because in leaf cells, glucose is the precursor of the antioxidant, L-ascorbate, and glucose-6-phosphate is a source of NADPH needed to support antioxidant capacity. A further objective was to establish whether the response of increased levels of glucose, sucrose, starch and G6P in leaves could be correlated with a greater degree of plant tolerance to O3. Four commercially available Spinacia oleracea varieties were screened for tolerance or susceptibility to detrimental effects of O3 employing one 6.5 hour acute exposure to 25O nL O3 L-1 air during the light. One day after the termination of ozonation (29 d post emergence), leaves of the plants were monitored both for damage and for gas exchange characteristics. Cultivar Winter Bloomsdale (cv Winter) leaves were least damaged on a quantitative grading scale. The leaves of cv Nordic, the most susceptible, were approximately 2.5 times more damaged. Photosynthesis (Pn) rates in the ozonated mature leaves of cv Winter were 48.9% less, and in cv Nordic, 66.2% less than in comparable leaves of their non-ozonated controls. Stomatal conductance of leaves of ozonated plants was found not to be a factor in the lower Pn rates in the ozonated plants. At some time points in the light, leaves of ozonated cv Winter plants had significantly higher levels of glucose, sucrose, starch, G6P, G1P, pyruvate and malate than did leaves of ozonated cv Nordic plants. It was concluded that leaves of cv Winter displayed a higher tolerance to ozone mediated stress than those of cv Nordic, in part because they had higher levels of glucose and G6P that could be mobilized during diminished photosynthesis to generate antioxidants (e.g., ascorbate) and reductants (e.g., NADPH). Elevated levels of both pyruvate and malate in the leaves of ozonated cv Winter suggested an increased availability of respiratory substrates to support higher respiratory capacity needed for repair, growth, and maintenance.Abbreviations ADPG-PPiase
ADPglucose pyrophosphorylase
- ASC
L-ascorbic acid
- APX
ascorbate peroxidase
- Ce
CO2 concentration in air in the measuring cuvette during photosynthesis measurements
- Ci
CO2 concentration in the leaf intercellular spaces during photosynthesis measurement
- Chl
chlorophyll
- DHA
dehydroascorbic acid
- DHA reductase
dehydroascorbate reductase
- DHAP
dihydroxyacetone phosphate
- GAP
glyceraldehyde-3-phosphate
- Gluc
glucose
- GR
glutathione reductase
- Gsw
stomatal conductance with units as mmol H2O m-2 s-1
- GSSG
oxidized glutathione
- GSH
reduced glutathione
- G1P
glucose-1-phosphate
- G6P
glucose-6-phosphate
- G6P dehydrogenase
glucose-6-phosphate dehydrogenase
- 6PG
6-phosphogluconate
- 6PG dehydrogenase
6-phosphogluconate dehydrogenase
- F6P
fructose-6-phosphate
- FBP
fructose-1,6-bisphosphate
- MAL
malate
- MDHA reductase
monodehydroascorbate reductase
- PE
post-emergence
- PEP
phosphoenolpyruvate
- PGA
3-phosphoglycerate
- Pi
orthophosphate
- PYR
pyruvate
- Pn
net CO2 photoas-similation in leaves
- PPFD
photosynthetic photon flux density with units of mol photons m-2 s-1
- PPRC
pentose phosphate reductive cycle
- RuBP
ribulose-1,5-bisphosphate
- rubisco
ribulose-1,5-bisphosphate carboxylase/oxygenase
- SLW
specific leaf weight
- TCA cycle
tricarboxylic acid cycle
- Triose-P
DHAP+GAP 相似文献
3.
Leaves on transgenic tobacco plants expressing yeast-derived invertase in the apoplast develop clearly demarcated green and bleached sectors when they mature. The green areas contain low levels of soluble sugars and starch which are turned over on a daily basis, and have high rates of photosynthesis and low rates of respiration. The pale areas accumulate carbohydrate, photosynthesis is inhibited, and respiration increases. This provides a model system to investigate the sink regulation of photosynthetic metabolism by accumulating carbohydrate. The inhibition of photosynthesis is accompanied by a decrease of ribulose-1,5-bisphosphate and glycerate-3-phosphate, and an increase of triosephosphate and fructose-1,6-bisphosphate. The extracted activities of ribulose-1,5-bisphosphate carboxylase, fructose-1, 6-bisphosphatase and NADP-glyeraldehyde-3-phosphate dehydrogenase decreased. The activity of sucrose-phosphate synthase remained high or increased, an increased portion of the photosynthate was partitioned into soluble sugars rather than starch, and the pale areas showed few or no oscillations during transitions between darkness and saturating light in saturating CO2. The increased rate of respiration was accompanied by an increased level of hexose-phosphates, triose-phosphates and fructose-1,6-bisphosphate while glycerate-3-phosphate and phosphoenolpyruvate decreased and pyruvate increased. The activities of pyruvate kinase, phosphofructokinase and pyrophosphate: fructose-6-phosphate phosphotransferase increased two- to four-fold. We conclude that an increased level of carbohydrate leads to a decreased level of Calvin-cycle enzymes and, thence, to an inhibition of photosynthesis. It also leads to an increased level of glycolytic enzymes and, thence, to a stimulation of respiration. These changes of enzymes are more important in middle- or long-term adjustments to high carbohydrate levels in the leaf than fine regulation due to depletion of inorganic phosphate or high levels of phosphorylated metabolites.Abbreviations Fru 1,6bisP
fructose-1,6-bisphosphate
- Fru 1,6bisPase
fructose-1,6-bisphosphatase
- Fru6P
fructose-6-phosphate
- Glc 1P
glucose-1-phosphate
- Glc6P
glucose-6-phosphate
- NADP-GAPDH
NADP-dependent glyceraldehyde-3-phosphate dehydrogenase
- PFK
phosphofructokinase
- PEP
phosphoenolpyruvate
- PFP
pyrophosphate:fructose-6-phosphate phosphotransferase
- PGA
glycerate-3-phosphate
- PK
pyruvate kinase
- Pi
inorganic phosphate
- Ru1,5bisP
ribulose-1,5-bisphosphate
- Rubisco
ribulose-1,5-bisphosphate carboxylase-oxygenase
- SPS
sucrose-phosphate synthase
- triose-P
triose-phosphates 相似文献
4.
The maximum extractable activities of twenty-one photosynthetic and glycolytic enzymes were measured in mature leaves of Mesembryanthemum crystallinum plants, grown under a 12 h light 12 h dark photoperiod, exhibiting photosynthetic characteristics of either a C3 or a Crassulacean acid metabolism (CAM) plant. Following the change from C3 photosynthesis to CAM in response to an increase in the salinity of in the rooting medium from 100 mM to 400 mM NaCl, the activity of phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31) increased about 45-fold and the activities of NADP malic enzyme (EC 1.1.1.40) and NAD malic enzyme (EC 1.1.1.38) increased about 4- to 10-fold. Pyruvate, Pi dikinase (EC 2.7.9.1) was not detected in the non-CAM tissue but was present in the CAM tissue; PEP carboxykinase (EC 4.1.1.32) was detected in neither tissue. The induction of CAM was also accompanied by large increases in the activities of the glycolytic enzymes enolase (EC 4.2.1.11), phosphoglyceromutase (EC 2.7.5.3), phosphoglycerate kinase (EC 2.7.2.3), NAD glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12), and glucosephosphate isomerase (EC 2.6.1.2). There were 1.5- to 2-fold increases in the activities of NAD malate dehydrogenase (EC 1.1.1.37), alanine and aspartate aminotransferases (EC 2.6.1.2 and 2.6.1.1 respectively) and NADP glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.13). The activities of ribulose-1,5-bisphosphate (RuBP) carboxylase (EC 4.1.1.39), fructose-1,6-bisphosphatase (EC 3.1.3.11), phosphofructokinase (EC 2.7.1.11), hexokinase (EC 2.7.1.2) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) remained relatively constant. NADP malate dehydrogenase (EC 1.1.1.82) activity exhibited two pH optima in the non-CAM tissue, one at pH 6.0 and a second at pH 8.0. The activity at pH 8.0 increased as CAM was induced. With the exceptions of hexokinase and glucose-6-phosphate dehydrogenase, the activities of all enzymes examined in extracts from M. crystallinum exhibiting CAM were equal to, or greater than, those required to sustain the maximum rates of carbon flow during acidification and deacidification observed in vivo. There was no day-night variation in the maximum extractable activities of phosphoenolpyruvate carboxylase, NADP malic enzyme, NAD malic enzyme, fructose-1,6-bisphosphatase and NADP malate dehydrogenase in leaves of M. crystallinum undergoing CAM.Abbreviations CAM
Crassulacean acid metabolism
- PEP
phosphoenolpyruvate
- RuBP
ribulose-1,5-bisphosphate 相似文献
5.
Liver metabolism in cold hypoxia: a comparison of energy metabolism and glycolysis in cold-sensitive and cold-resistant mammals 总被引:2,自引:0,他引:2
T. A. Churchill K. M. Cheetham S. Simpkin C. J. Green L. C. H. Wang B. J. Fuller 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1994,164(5):396-404
The effects of cold hypoxia were examined during a time-course at 2 °C on levels of glycolytic metabolites: glycogen, glucose, glucose-1-phosphate, glucose-6-phosphate, fructose-6-phosphate, fructose-1,6-bisphosphate, phosphoenolpyruvate, pyruvate, lactate and energetics (ATP, ADP, AMP) of livers from rats and columbian ground squirrels. Responses of adenylate pools reflected the energy imbalance created during cold hypoxia in both rat and ground squirrel liver within minutes of organ isolation. In rat, ATP levels and energy charge values for freshly isolated livers were 2.54 mol·g-1 and 0.70, respectively. Within 5 min of cold hypoxia, ATP levels had dropped well below control values and by 8 h storage, ATP, AMP, and energy charge values were 0.21 mol·g-1, 2.01 mol·g-1, and 0.17, respectively. In columbian ground squirrels the patterns of rapid ATP depletion and AMP accumulation were similar to those found in rat. In rat liver, enzymatic regulatory control of glycolysis appeared to be extremely sensitive to the decline in cellular energy levels. After 8 h cold hypoxia levels of fructose-6-phosphate decreased and fructose-1,6-bisphosphate increased, thus reflecting an activation of glycolysis at the regulatory step catalysed by phospho-fructokinase fructose-1,6-bisphosphatase. Despite an initial increase in flux through glycolysis over the first 2 min (lactate levels increased 3.7 mol·g-1), further flux through the pathway was not permitted even though glycolysis was activated at the phosphofructokinase/fructose-1,6-bisphosphatase locus at 8 h, since supplies of phosphorylated substrate glucose-1-phosphate or glucose-6-phosphate remained low throughout the duration of the 24-h period. Conversely, livers of Columbian ground squirrels exhibited no activation or inactivation of two key glycolytic regulatory loci, phosphofructokinase/fructose-1,6-bisphosphatase and pyruvate kinase/phosphoenolpyruvate carboxykinase and pyruvate carboxylase. Although previous studies have shown similar allosteric sensitivities to adenylates to rat liver phospho-fructokinase, there was no evidence of an activation of the pathway as a result of decreasing high energy adenylate, ATP or increasing AMP levels. The lack of any apparent regulatory control of glycosis during cold hypoxia may be related to hibernator-specific metabolic adaptations that are key to the survival of hypothermia during natural bouts of hibernation.Abbreviations DHAP
dihydroxyacetonephosphate
- EC
energy charge
- F1,6P2
fructose-1,6-bisphosphate
- F2,6P2
fructose-2,6-bisphosphate
- F6P
fructose-6-phosphate
- FBP
fructose-1,6-bisphosphatase
- G1P
glucose-1-phosphate
- G6P
glucose-6-phosphate
- GAP
glyceraldehyde-3-phosphate
- GAPDH
glyceraldehyde-3-phosphate dehydrogenase
- L/R
lactobionate/raffinose-based solution
- MR
metabolic rate
- PDH
pyruvate dehydrogenase
- PEP
phosphoenolpyruvate
- PEPCK & PC
phosphoenolpyruvate carboxykinase and pyruvate carboxylase
- PFK
phosphofructokinase; PK, pyruvate kinase
-
Q
10
the effect of a 10 °C drop in temperature on reaction rates (generally, Q
10=2–3)
- TA
total adenylates
- UW solution
University of Wisconsin solution (L/R-based) 相似文献
6.
Regulation of sucrose-starch accumulation and its effect on CO2 gas exchange and electron transport were studied in low-temperature-stressed and cold-acclimated spring (Katepwa) and winter (Monopol) cultivars of wheat (Triticum aestivum L.). Low-temperature stress of either the spring or winter cultivar was associated with feedback-limited photosynthesis as indicated by a 50–60% reduction in CO2 assimilation rates, twofold lower ATP/ADP ratio, and threefold lower electron transport rate than 20°C-grown control plants. However, no limitations were evident at the level of ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco) in low-temperature-stressed plants. Cold acclimation of the spring cultivar resulted in similar feedback-limited photosynthesis observed during low-temperature stress. In contrast, cold acclimation of the winter cultivar resulted in an adjustment of CO2 assimilation rates to that of control plants. However, we show, for the first time, that this capacity to adjust CO2 assimilation still appeared to be associated with limited triose phosphate utilisation, a twofold lower ATP/ADP ratio, a reduction in electron transport rates but no restriction at the level of Rubisco compared to controls grown at 20°C. Thus, contrary to previous suggestions, we conclude that cold-acclimated Monopol appears to exhibit feedback limitations at the level of electron transport characteristic of cold-stressed plants despite the maintenance of high rates of CO2 assimilation. Furthermore, the differential capacity of the winter cultivar to adjust CO2 assimilation rates was associated with higher levels of sucrose accumulation and a threefold higher sucrose-phosphate synthase activity despite an apparent limitation in triose phosphate utilisation.Abbreviations AGPase
ADP-glucose pyrophosphorylase
- FBPase
fructose-1,6-bisphosphatase
- Fru 6-P
fructose 6-phosphate
- Fru 1,6-BP
fructose 1,6-bisphosphate
- Glc 6-P
glucose 6-phosphate
- PGA
3-phosphoglyceric acid
- Rubisco
ribulose-1,5-bisphosphate carboxylase-oxygenase
- RuBP
ribulose 1,5-bisphosphate
- SPS
sucrose-phosphate synthase
- Triose-P
triose phosphate 相似文献
7.
The metabolite levels in the mesophyll of leaves of Zea mays L. have been compared with the regulatory properties of the cytosolic fructose-1,6-bisphosphatase from the mesophyll to show how withdrawal of triose phosphate for sucrose synthesis is reconciled with generation of the high concentrations of triose phosphate which are needed to allow intercellular diffusion of carbon during photosynthesis. i) A new technique is presented for measuring the intercellular distribution of metabolites in maize. The bundle-sheath and mesophyll tissues are partially separated by differential homogenization and filtration through nylon nets under liquid nitrogen. ii) considerable gradients of 3-phosphoglycerate, triose phosphate, malate and phosphoenolpyruvate exist between the mesophyll and bundle sheath which would allow intercellular shuttles to be driven by diffusion. These gradients could result from the distribution of electron transport and the Calvin cycle in maize leaves. iii) consequently, the mesophyll contains high concentrations of triose phosphate and fructose-1,6-bisphosphate. iv) Most of the regulator metabolite fructose-2,6-bisphosphate, is present in the mesophyll. v) The cytosolic fructose-1,6-bisphosphatase has a lower substrate affinity than that found for the enzyme from C3 species, especially in the presence of inhibitors like fructose-2,6-bisphosphate. vi) This lowered affinity for substrate makes it possible to reconcile use of triose phosphate for sucrose synthesis with the maintenance of the high concentration of triose phosphate in the mesophyll needed for operation of photosynthesis in this species.Abbreviations DHAP
Dihydroxyacetonephosphate
- Fru1,6-bisP
fructose-1,6-bisphosphate
- Fru2,6bisP
fructose-2,6-bisphosphate
- PEP(Case)
phosphoenolpyruvate (carboxylase)
- PGA
3-phosphoglycerate
- Rubisco
ribulose-1,5-bisphosphate carboxylase 相似文献
8.
The relationship between the gas-exchange characteristics of attached leaves of Amaranthus edulis L. and the contents of photosynthetic intermediates was examined in response to changing irradiance and intercellular partial pressure of CO2. After determination of the rate of CO2 assimilation at known intercellular CO2 pressure and irradiance, the leaf was freeze-clamped and the contents of ribulose-1,5-bisphosphate, glycerate-3-phosphate, fructose-1,6-bisphosphate, glucose-6-phosphate, fructose-6-phosphate, triose phosphates, phosphoenolpyruvate, pyruvate, oxaloacetate, aspartate, alanine, malate and glutamate were measured. A comparison between the sizes of metabolite pools and theoretical calculations of metabolite gradients required for transport between the mesophyll and the bundle-sheath cells showed that aspartate, alanine, glycerate-3-phosphate and triose phosphates were present in sufficient quantities to support transport by diffusion, whereas pyruvate and oxaloacetate were not likely to contribute appreciably to the flux of carbon between the two cell types. The amounts of ribulose-1,5-bisphosphate were high at low intercellular partial pressures of CO2, and fell rapidly as the CO2-assimilation rate increased with increasing intercellular partial pressures of CO2, indicating that bundle-sheath CO2 concentrations fell at low intercellular partial pressures of CO2. In contrast, the amount of phosphoenolpyruvate and of C4-cycle intermediates declined at low intercellular partial pressures of CO2. This behaviour is discussed in relation to the co-ordination of carbon assimilation between the Calvin and C4 cycles.Abbreviations PEP
phosphoenolpyruvate
- PGA
glycerate-3-phosphate
-
p
i
intercellular CO2 pressure
- RuBP
ribulose-1,5-bisphosphate
- triose-P
triose phosphates 相似文献
9.
Mohammad Hajirezaei Uwe Sonnewald Roberto Viola Sarah Carlisle David Dennis Mark Stitt 《Planta》1993,192(1):16-30
Potato (Solanum tuberosum L.) plants were transformed with antisense constructs to the genes encoding the -and -subunits of pyrophosphate: fructose-6-phosphate phosphotransferase (PEP), their expression being driven by the constitutive CaMV 35S promotor. (i) In several independent transformant lines, PFP expression was decreased by 70–90% in growing tubers and by 88–99% in stored tubers. (ii) The plants did not show any visual phenotype, reduction of growth or decrease in total tuber yield. However, the tubers contained 20–40% less starch than the wild type. Sucrose levels were slightly increased in growing tubers, but not at other stages. The rates of accumulation of sucrose and free hexoses when tubers were stored at 4° C and the final amount accumulated were the same in antisense and wild-type tubers. (iii) Metabolites were investigated at four different stages in tuber life history; growing (sink) tubers, mature tubers, cold-sweetening tubers and sprouting (source) tubers. At all stages, compared to the wild type, antisense tubers contained slightly more hexose-phosphates, two- to threefold less glycerate-3-phosphate and phosphoenolpyruvate and up to four-to fivefold more fructose-2,6-bisphosphate. (iv) There was no accumulation or depletion of inorganic pyrophosphate (PPi), or of UDP-glucose relative to the hexose-phosphates. (v) The pyruvate content was unaltered or only marginally decreased, and the ATP/ADP ratio did not change. (vi) Labelling experiments on intact tubers did not reveal any significant decrease in the unidirectional rate of metabolism of [U-14C]sucrose to starch, organic acids or amino acids. Stored tubers with an extreme (90%) reduction of PFP showed a 25% decrease in the metabolism of [U14-C] sucrose. (vii) Metabolism (cycling) of [U-14C]glucose to surcrose increased 15-fold in discs from growing antisense tubers, compared with growing wild-type tubers. Resynthesis of sucrose was increased by 10–20% when discs from antisense and wild-type tubers stored at 4° C (cold sweetening) were compared. The conversion of [U-14C]glucose to starch was decreased by about 30% and 50%, respectively. (viii) The randomisation of [1-13C]glucose in the glucosyl and fructosyl moieties of sucrose was decreased from 13.8 and 15.7% in the wild type to 3.6 and 3.9% in an antisense transformant. Simultaneously, randomisation in glucosyl residues isolated from starch was reduced from 14.4 to 4.1%. (ix) These results provide evidence that PFP catalyses a readily reversible reaction in tubers, which is responsible for the recycling of label from triose-phosphates to hexose-phosphates, but with the net reaction in the glycolytic direction. The results do not support the notion that PFP is involved in regulating the cytosolic PPi concentration. They also demonstrate that PFP does not control the rate of glycolysis, and that tubers contain exessive capacity to phosphorylate fructose-6-phosphate. The decreased concentration of phosphoenolpyruvate and glycerate-3-phosphate compensates for the decrease of PFP protein by stimulating ATP-dependent phosphofructokinase, and by stimulating fructose-6-phosphate,2-kinase to increase the fructose-2,6-bisphosphate concentration and activate the residual PFP. The decreased starch accumulation is explained as an indirect effect, caused by the increased rate of resynthesis (cycling) of sucrose in the antisense tubers.Abbreviations Fru1,6bisP
fructose-1,6-bisphosphate
- Fru2,6bisP
fructose-2,6-bisphosphate
- Fru6P
fructose-6-phosphate
- Glc1P
glucose-1-phosphate
- Glc6P
glucose-6-phosphate
- NMR
nuclear magnetic resonance
- 3PGA
glycerate-3-phosphate
- PEP
phosphoenolpyruvate
- PEP
pyrophosphate: fructose-6-phosphate phosphotransferase
- PFK
phosphofructokinase
- UDPGlc
UDP glucose
- WT
wild type
This research was supported by the Bundesministerium for Forschung and Technology (M.S., U.S.), the Canadian Research Council (S.C., D.D.), the Agricultural and Food Research Council (R.V.) and Sandoz Agro Ltd. (M.H., M.S.). 相似文献
10.
The class II fructose-1,6-bisphosphatase gene of Corynebacterium glutamicum, fbp, was cloned and expressed with a N-terminal His-tag in Escherichia coli. Purified, His-tagged fructose-1,6-bisphosphatase from C. glutamicum was shown to be tetrameric, with a molecular mass of about 140 kDa for the homotetramer. The enzyme displayed Michaelis-Menten kinetics for the substrate fructose 1,6-bisphosphate with a Km value of about 14 µM and a Vmax of about 5.4 µmol min–1 mg–1 and kcat of about 3.2 s–1. Fructose-1,6-bisphosphatase activity was dependent on the divalent cations Mg2+ or Mn2+ and was inhibited by the monovalent cation Li+ with an inhibition constant of 140 µM. Fructose 6-phosphate, glycerol 3-phosphate, ribulose 1,5-bisphosphate and myo-inositol-monophosphate were not significant substrates of fructose-1,6-bisphosphatase from C. glutamicum. The enzymatic activity was inhibited by AMP and phosphoenolpyruvate and to a lesser extent by phosphate, fructose 6-phosphate, fructose 2,6-bisphosphate, and UDP. Fructose-1,6-bisphosphatase activities and protein levels varied little with respect to the carbon source. Deletion of the chromosomal fbp gene led to the absence of any detectable fructose-1,6-bisphosphatase activity in crude extracts of C. glutamicum WTfbp and to an inability of this strain to grow on the carbon sources acetate, citrate, glutamate, and lactate. Thus, fbp is essential for growth on gluconeogenic carbon sources and likely codes for the only fructose-1,6-bisphosphatase in C. glutamicum. 相似文献
11.
Phyllodia of the Crassulacean acid metabolism (CAM) plant Kalanchoë tubiflora were allowed to fix 13CO2 in light and darkness during phase IV of the diurnal CAM cycle, and during prolongation of the regular light period. After 13CO2 fixation in darkness, only singly labelled [13C]malate molecules were found. Fixation of 13CO2 under illumination, however, produced singly labelled malate as well as malate molecules which carried label in two, three or four carbon atoms. When the irradiance during 13CO2 fixation was increased, the proportion of singly labelled malate decreased in favour of plurally labelled malate. The irradiance, however, did not change either the ratio of labelled to unlabelled malate molecules found in the tissue after the 13CO2 application, or the magnitude of malate accumulation during the treatment with label. The ability of the tissue to store malate and the labelling pattern changed throughout the duration of the prolonged light period. The results indicate that malate synthesis by CAM plants in light can proceed via a pathway containing two carboxylation steps, namely ribulose-1,5-bisphosphate-carboxylase/oxygenase (EC 4.1.1.39) and phosphoenolpyruvate carboxylase (EC 4.1.1.31) which operate in series and share common intermediates. It can be concluded that, in light, phosphoenolpyruvate carboxylase can also synthesize malate independently of the proceeding carboxylation step by ribulose-1,5-bisphosphate carboxylase/oxygenase.Abbreviations CAM
Crassulacean acid metabolism
- PEP
phosphoenolpyruvate
- PEPCase
phosphoenolpyruvate carboxylase (EC 4.1.1.31)
- RuBPCase
ribulose-1,5-bisphosphate carboxylase/oxygenase (EC 4.1.1.39)
- TMS
trimethylsilyl 相似文献
12.
Thomas A. Churchill Kenneth B. Storey 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1989,159(4):461-472
Summary The course of glycerol biosynthesis, initiated by exposure to –4°C, was monitored in larvae of the goldenrod gall moth,Epiblema scudderiana, and accompanying changes in the levels of intermediates of glycolysis, adenylates, glycogen, glucose, fructose-2,6-bisphosphate, and fermentative end products were characterized. Production of cryoprotectant was initiated within 6 h after a switch from +16° to –4°C, with halfmaximal levels reached in 30 h and maximal content, 450–500 mol/g wet weight, achieved after 4 days. Changes in the levels of intermediates of the synthetic pathway within 2 h at –4°C indicated that the regulatory sites involved glycogen phosphorylase, phosphofructokinase, and glycerol-3-phosphatase. A rapid increase in fructose-2,6-bisphosphate, an activator of phosphofructokinase and inhibitor of fructose-1,6-bisphosphatase, appeared to have a role in maintaining flux in the direction of glycerol biosynthesis. Analysis of metabolite changes as glycerol production slowed suggested that the inhibitory restriction of the regulatory enzymes was slightly out of phase. Inhibition at the glycerol-3-phosphatase locus apparently occurred first and resulted in a build-up of glycolytic intermediates and an overflow accumulation of glucose. Glucose inhibition of phosphorylase, stimulating the conversion of the activea to the inactiveb forms, appears to be the mechanism that shuts off phosphorylase function, counteracting the effects of low temperature that are the basis of the initial enzyme activation. Equivalent experiments carried out under a nitrogen gas atmosphere suggested that the metabolic make-up of the larvae in autumn is one that obligately routes carbohydrate flux through the hexose monophosphate shunt. The consequence of this is that fermentative ATP production during anoxia is linked to the accumulation of large amounts of glycerol as the only means of maintaining redox balance.Abbreviations
G6P
glucose-6-phosphate
-
F6P
fructose-6-phosphate
-
F1, 6P
fructose-1,6-bisphosphate
-
F2,6P
2
fructose-2,6-bisphosphate
-
G3P
grycerol-3-phosphate
-
DHAP
dinydroxyacetonephosphate
-
GAP
glyceraldehyde-3-phosphate
-
PEP
phosphoenolpyruvate
-
PFK
phosphofructokinase
-
FBPase
fructose-1,6-bisphosphatase
-
PK
pyruvate kinase 相似文献
13.
Glycolytic Flux Is Adjusted to Nitrogenase Activity in Nodules of
Detopped and Argon-Treated Alfalfa Plants 总被引:4,自引:1,他引:3
Paola M.G. Curioni Ueli A. Hartwig Josef N?sberger Kathryn A. Schuller 《Plant physiology》1999,119(2):445-454
To investigate the short-term (30–240 min) interactions among nitrogenase activity, NH4+ assimilation, and plant glycolysis, we measured the concentrations of selected C and N metabolites in alfalfa (Medicago sativa L.) root nodules after detopping and during continuous exposure of the nodulated roots to Ar:O2 (80:20, v/v). Both treatments caused an increase in the ratios of glucose-6-phosphate to fructose-1,6-bisphosphate, fructose-6-phosphate to fructose-1,6-bisphosphate, phosphoenolpyruvate (PEP) to pyruvate, and PEP to malate. This suggested that glycolytic flux was inhibited at the steps catalyzed by phosphofructokinase, pyruvate kinase, and PEP carboxylase. In the Ar:O2-treated plants the apparent inhibition of glycolytic flux was reversible, whereas in the detopped plants it was not. In both groups of plants the apparent inhibition of glycolytic flux was delayed relative to the decline in nitrogenase activity. The decline in nitrogenase activity was followed by a dramatic increase in the nodular glutamate to glutamine ratio. In the detopped plants this was coincident with the apparent inhibition of glycolytic flux, whereas in the Ar:O2-treated plants it preceded the apparent inhibition of glycolytic flux. We propose that the increase in the nodular glutamate to glutamine ratio, which occurs as a result of the decline in nitrogenase activity, may act as a signal to decrease plant glycolytic flux in legume root nodules. 相似文献
14.
The aim of this work was to identify which aspects of photosynthetic metabolism respond most sensitively to leaf water deficit. Spinach (Spinacia oleracea L.) leaf discs were floated on sorbitol concentrations of increasing molarity and changes of the protoplast volume were estimated using [14C]sorbitol and 3H2O penetration. Detached leaves were also wilted until 10% of their fresh weight was lost. Photosynthesis was studied at very high external CO2 concentrations, to eliminate the effect of closing stomata. There was no large inhibition of CO2 fixation after wilting leaves, or until the external water deficit was greater than-1.2 MPa. However, partitioning changed markedly at these moderate water deficits: more sucrose and less starch was made. When an inhibition of CO2-saturated photosynthesis did appear at a water deficit of-2.0 MPa and above, measurements of chlorophyll-fluorescence quenching and metabolite levels showed the thylakoid reactions were not especially susceptible to short-term water stress. The inhibition was accompanied by a small increase of the triose phosphate: ribulose-1,5-bisphosphate ratio, showing regeneration of ribulose-1,5-bisphosphate was affected. However, there was also a general increase of the estimated concentrations of most metabolites, indicating that there is no specific site for the inhibition of photosynthesis. Increasing water deficit led to a large increase of fructose-2,6-bisphosphate. This is explained in terms of a simultaneous increase of fructose-6-phosphate and inorganic phosphate as the cell shrinks. The high fructose-2,6-bisphosphate led to the accumulation of triose phosphates, and the potential significance of this for protection against photoinhibition is discussed. There was an increase in the extractable activity of sucrose-phosphate synthase. This was only detected when the enzyme was assayed in conditions which distinguish between different kinetic forms which have previously been identified in spinach leaves. It is proposed that activation of sucrose-phosphate synthase is one of the first sites at which spinach leaves respond to a rising water deficit. This could be of importance for osmoregulation.Abbreviations Chl
chlorophyll
- Fru1,6bisP
fructose-1,6-bisphosphate
- Fru2,6bisP
fructose-2,6-bisphosphate
- Fru6P
fructose-6-phosphate
- Glc6P
glucose-6-phosphate
- PGA
glycerate-3-phosphate
- Pi
inorgamic phosphate
- Ru1,5bisP
ribulose-1,5-bisphosphate
- SPS
sucrose-phosphate synthase
- triose-P
sum of glyceraldehyde-3-phosphate and dehydroxyacetone phosphate
- UDPGlc
uridine diphosphoglucose 相似文献
15.
Spinach leaf discs were floated on methyl-viologen solutions (5–200 nmol·l-1) and the effect on photosynthetic metabolism was then investigated under conditions of saturating CO2. Methyl viologen led to increased non-photochemical quenching, and the ATP/ADP ratio increased from <2 to >10. Comparison of the apparent quantum yield and non-photochemical quenching indicated that these concentrations of methyl viologen were only catalysing a marginal electron flux, and that the decrease in quantum yield was mainly the result of pH-triggered energy dissipation. Similar changes were also obtained after supplying tentoxin to inhibit the chloroplast ATP synthase and increase the energisation of the thylakoids. The photosystem-II acceptor, QA, was monitored by photochemical fluorescence quenching, and became more reduced. In contrast, the activation of NADP-malate dehydrogenase decreased, showing that the acceptor side of photosystem I becomes more oxidised. Similar changes were observed after supplying tentoxin. It is concluded that increased thylakoid energisation can lead to a substantial restriction of linear electron transport. Analysis of metabolite levels showed that glycerate-3-phosphate reduction was imporved, but that there was a large accumulation of triose phosphates and fructose-1,6-bisphosphate. This is the consequence of an inhibition of the regeneration of ribulose-1,5-bisphosphate, caused by inactivation of the stromal fructose-1,6-bisphosphatase and, to a lesser extent, phosphoribulokinase. Methyl viologen also led to inactivation of sucrose-phosphate synthase, and abolished the response of fructose-2,6-bisphosphate to rising rates of photosynthesis. This provides evidence for a primary role of glycerate-3-phosphate in controlling the activity of fructose-6-phosphate, 2-kinase and, thence, the fructose-2,6-bisphosphate concentration as the rate of photosynthesis increases. It is concluded that the very moderate ATP/ADP ratios found in chloroplasts are the results of constraints on the operation of ATP synthase. They can be increased if the thylakoid energisation is increased. However, the increased energisation acts directly or indirectly to disrupt many other aspects of photosynthetic metabolism including linear electron transport, activation of the Calvin cycle, and the control of sucrose and starch synthesis.Abbreviations and symbols Frul,6P2 (Fru1,6Pase)
fructose-1,6-bisphosphate(ase)
- Fru2,6P, (Fru2,6Pase)
fructose-2,6-bisphosphate(-ase)
- Fru6P
fructose-6-phosphate
- Glc6P
glucose-6-phosphate
- Pi
inorganic phosphate
- PSI and PSII
photosystems I and II
- qE
high energy' quenching of chlorophyll fluorescence
- PGA
glycerate-3-phosphate
- QA
primary stable acceptor of PSII
- Ru5P (Ru1,5P2)
ribulose-5-phosphate (-1,5-bisphosphate)
- SPS
sucrose-phosphate synthase
- triose P
dihydroxyacetone phosphate plus glyceraldehyde-3-phosphate
-
s
apparent quantum yield
Dedicated to Professor E. Latzko on the occasion of his 65th birthday 相似文献
16.
Differential inhibition of photosynthesis during pre-flowering drought stress in Sorghum bicolor genotypes with different senescence traits 总被引:2,自引:0,他引:2
Young (16-day-old) Sorghum bicolor plants of a late- and slow-senescing Texas A&M line (B 35) and of an early- and fast-senescing descendant of an Ethiopian landrace (E 36-1) were subjected to drought stress by decreasing the soil water content to 30% field capacity over 6 days. Plant water potentials decreased from − 2 bar (controls) to − 10 to − 18 bar, and this drought stress resulted in: (1) differential phenotypic reactions and (2) differential decreases in photosynthesis rates in the two cultivars. While E 36-1 tended to lose viable leaf area from the leaf tips downwards, B 35 showed a gradual overall drying of the leaf. At the same time, photosynthesis rates decreased from 31.5 ± 1.6 to 12.3 ± 5.0 µmol CO2 m−2 s−1 (E 36-1) and from 30.5 ± 1.6 to 3.3 ± 2.6 µmol CO2 m−2 s−1 (B 35), respectively. In vitro enzyme activities of phosphoenolpyruvate carboxylase (PEPCase), malate dehydrogenase (MDH) and malic enzyme (ME) on a leaf area basis exceeded the photosynthesis rates. Pyruvate phosphate dikinase (PPDK) activity was close to the photosynthesis rates in control plants and higher than the photosynthesis rates in drought-stressed plants. Thus, none of the enzymes appeared to limit photosynthesis under drought stress, and likely bottleneck enzyme activities of the C3 pathway in the bundle-sheath cells, i.e. ribulose-1,5-bisphosphate carboxylase (RubisCO) and stromal fructose-1,5-bisphosphatase (sFBPase), also showed sufficient activities to sustain higher photosynthesis rates than those observed in the stressed plants. However, under drought stress, total leaf malate concentrations were higher in B 35 (up to 33.1 µmol g−1 FW) than in E 36-1 (up to 22.4 µmol g−1 FW). In particular, at the presumed cytosolic pH of 7.0–7.3, S. bicolor PEPCase was strongly inhibited by malate. In contrast with the in vitro PEPCase enzyme activities, the A/Ci curves suggested a stronger decrease in the in vivo activity of the enzyme in B 35 under drought stress than in E 36-1. It is therefore suggested that photosynthesis under drought stress may be inhibited differentially through feedback malate inhibition of PEPCase in S. bicolor. 相似文献
17.
Light- and CO2-saturated photosynthesis of nonhardened rye (Secale cereale L. cv. Musketeer) was reduced from 18.10 to 7.17 mol O2·m–2·s–1 when leaves were transferred from 20 to 5°C for 30 min. Following cold-hardening at 5°C for ten weeks, photosynthesis recovered to 15.05 mol O2·m–2·s–1,comparable to the nonhardened rate at 20°C. Recovery of photosynthesis was associated with increases in the total activity and activation of enzymes of the photosynthetic carbon-reduction cycle and of sucrose synthesis. The total hexose-phosphate pool increase by 30% and 120% for nonhardened and cold-hardened leaves respectively when measured at 5°C. The large increase in esterified phosphate in coldhardened leaves occurred without a limitation in inorganic phosphate supply. In contrast, the much smaller increase in esterified phosphate in nonhardened leaves was associated with an inhibition of ribulose-1,5-bisphosphate carboxylase/oxygenase and sucrose-phosphate synthase activation. It is suggested that the large increases in hexose phosphates in cold-hardened leaves compensates for the higher substrate threshold concentrations needed for enzyme activation at low temperatures. High substrate concentrations could also compensate for the kinetic limitations imposed by product inhibition from the accumulation of sucrose at 5°C. Nonhardened leaves appear to be unable to compensate in this fashion due to an inadequate supply of inorganic phosphate.Abbreviations DHAP
dihydroxyacetone phosphate
- Fru6P
fructose-6-phosphate
- Fru 1,6BP
fructose-1,6-bisphosphate
- Fru1,6BPase
fructose-1,6-bisphosphatase
- Glc6P
glucose-6-phosphate
- PGA
3-phosphoglycerate
- PPFD
photosynthetic photon flux density
- CH
cold-hardened rye grown at 5°C
- NH
nonhardened rye grown at 24°C
- Rubisco
ribulose-1,5-bisphosphate carboxylase/oxygenase
- RuBP
ribulose-1,5-bisphosphate
- SPS
sucrose-phosphate synthase
- UDPGlc
uridine 5-diphosphoglucose
This work was supported by operating grants from the Swedish Natural Sciences Research Council to G.Ö. and P.G. 相似文献
18.
Reduced Cytosolic Fructose-1,6-Bisphosphatase Activity Leads to Loss of O(2) Sensitivity in a Flaveria linearis Mutant 总被引:4,自引:4,他引:0
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The mutant plant of Flaveria linearis characterized by Brown et al. (Plant Physiol. 81: 212-215) was studied to determine the cause of the reduced sensitivity to O2. Analysis of CO2 assimilation metabolites of freeze clamped leaves revealed that both 3-phosphoglycerate and ribulose 1,5-bisphosphate were high in the mutant plant relative to F. linearis with normal O2 sensitivity. The kcat of ribulose-1,5-bisphosphate carboxylase (RuBPCase) was equal in all plant material tested (range 18-22 s−1) indicating that no tight binding inhibitor was present. The degree of RuBPCase carbamylation was reduced in the mutant plant relative to the wild-type plant. Since 3-phosphoglycerate was high in the mutant plant and photosynthesis did not exhibit properties associated with RuBPCase limitations, we believe that the decarbamylation of RuBPCase was a consequence of another lesion in photosynthesis. Fructose 1,6-bisphosphate and its precursors, such as the triose phosphates, were in high concentration in the mutant plant relative to the wild type. The concentrations of the product of the fructose 1,6-bisphosphatase reaction, fructose 6-phosphate, and its isomer, glucose 6-phosphate, were the same in both plants. We found that the mutant plant had up to 75% less cytosolic fructose 1,6-bisphosphatase activity than the wild type but comparable levels of stromal fructose 1,6-bisphosphatase. We conclude that the reduced fructose-1,6-bisphosphatase activity restricts the mutant plant's capacity for sucrose synthesis and this leads to reduced or reversed O2 sensitivity. 相似文献
19.
(i) We have studied the influence of reduced phosphoglucose-isomerase (PGI) activity on photosynthetic carbon metabolism in mutants of Clarkia xantiana Gray (Onagraceae). The mutants had reduced plastid (75% or 50% of wildtype) or reduced cytosolic (64%, 36% or 18% of wildtype) PGI activity. (ii) Reduced plastid PGI had no significant effect on metabolism in low light. In high light, starch synthesis decreased by 50%. There was no corresponding increase of sucrose synthesis. Instead glycerate-3-phosphate, ribulose-1,5-bisphosphate, reduction of QA (the acceptor for photosystem II) and energy-dependent chlorophyll-fluorescence quenching increased, and O2 evolution was inhibited by 25%. (iii) Decreased cytosolic PGI led to lower rates of sucrose synthesis, increased fructose-2,6-bisphosphate, glycerate-3-phosphate and ribulose-1,5-bisphosphate, and a stimulation of starch synthesis, but without a significant inhibition of O2 evolution. Partitioning was most affected in low light, while the metabolite levels changed more at saturating irradiances. (iv) These results provide decisive evidence that fructose-2,6-bisphosphate can mediate a feedback inhibition of sucrose synthesis in response to accumulating hexose phosphates. They also provide evidence that the ensuing stimulation of starch synthesis is due to activation of ADP-glucose pyrophosphorylase by a rising glycerate-3-phosphate: inorganic phosphate ratio, and that this can occur without any loss of photosynthetic rate. However the effectiveness of these mechanisms varies, depending on the conditions. (v) These results are analysed using the approach of Kacser and Burns (1973, Trends Biochem. Sci. 7, 1149–1161) to provide estimates for the elasticities and flux-control coefficient of the cytosolic fructose-1,6-bisphosphatase, and to estimate the gain in the fructose-2,6-bisphosphate regulator cycle during feedback inhibition of sucrose synthesis.Abbreviations and symbols Chl
chlorophyll
- Fru6P
fructose-6-phosphate
- Frul,6bisP
fructose-1,6-bisphosphate
- Fru-1,6Pase
fructose-1,6-bisphosphatase
- Fru2,6bisP
fructose-2,6-bisphosphate
- Fru2,6Pase
fructose-2,6-bisphosphatase
- Glc6P
glucose-6-phosphate
- PGI
phosphoglucose isomerase
- Pi
inorganic phosphate
- QA
acceptor for photosystem II
- Ru1,5bisP
ributose-1,5-bisphosphate
- SPS
sucrose-phosphate synthase 相似文献
20.
The hyperthermophilic archaeon Pyrococcus furiosus was grown on pyruvate as carbon and energy source. The enzymes involved in gluconeogenesis were investigated. The following findings indicate that glucose-6-phosphate formation from pyruvate involves phosphoenolpyruvate synthetase, enzymes of the Embden-Meyerhof pathway and fructose-1,6-bisphosphate phosphatase.Cell extracts of pyruvate-grown P.furiosus contained the following enzyme activities: phosphoenolpyruvate synthetase (0.025 U/mg, 50 °C), enolase (0.9 U/mg, 80 °C), phosphoglycerate mutase (0.13 U/mg, 55 °C), phosphoglycerate kinase (0.01 U/mg, 50 °C), glyceraldehyde-3-phosphate dehydrogenase reducing either NADP+ or NAD+ (NADP+: 0.019 U/mg, NAD+: 0.009 U/mg; 50 °C), triosephosphate isomerase (1.4 U/mg, 50 °C), fructose-1,6-bisphosphate aldolase (0.0045 U/mg, 55 °C), fructose-1,6-bisphosphate phosphatase (0.026 U/mg, 75 °C), and glucose-6-phosphate isomerase (0.22 U/mg, 50 °C). Kinetic properties (V
max values and apparent K
m values) of the enzymes indicate that they operate in the direction of sugar synthesis. The specific enzyme activities of phosphoglycerate kinase, glyceraldehyde-3-phosphate dehydrogenase (NADP+-reducing) and fructose-1,6-bisphosphate phosphatase in pyruvate-grown P. furiosus were by a factor of 3, 10 and 4, respectively, higher as compared to maltose-grown cells suggesting that these enzymes are induced under conditions of gluconeogenesis. Furthermore, cell extracts contained ferredoxin: NADP+ oxidoreductase (0.023 U/mg, 60 °C); phosphoenolpyruvate carboxylase (0.018 U/mg, 50 °C) acts as an anaplerotic enzyme.Thus, in P. furiosus sugar formation from pyruvate involves reactions of the Embden-Meyerhof pathway, whereas sugar degradation to pyruvate proceeds via a modified non-phosphorylated Entner-Doudoroff pathway. 相似文献