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1.
When purified acetyl-CoA carboxylase was incubated with various phospholipids, the effects on carboxylase activity were quite diverse. Phosphatidic acid, phosphatidylcholine, and phosphatidylinositol were slightly stimulatory, whereas carboxylase was inhibited by polyphosphoinositides in a time- and concentration-dependent manner. Phosphatidylinositol 4,5-bisphosphate (TPI) was the most effective inhibitor; carboxylase activity was inhibited 50% after incubation with 1.5 μm TPI for 30 min. Incubation of carboxylase with citrate reduced the susceptibility to inhibition by TPI. The inhibition was reversed by removal of TPI from the inhibited enzyme. Incubation of TPI with divalent metal cations removed its ability to inhibit carboxylase. Sedimentation studies showed that TPI treatment shifts carboxylase to a less-polymerized form. The Km for ATP, 24 μm, was not affected by the inhibitor. However, the apparent Km for acetyl-CoA was decreased from 44 to 11 μm following incubation with TPI. The possibility that polyphosphoinositides may play a role in acetyl-CoA carboxylase regulation is discussed. 相似文献
2.
《Bioorganic & medicinal chemistry》2019,27(22):115120
Ribulose-1,5-biphosphate carboxylase/oxygenase (RuBisCO), an enzyme in the Calvin-Benson-Bassham cycle of photosynthesis, catalyzes the first step of CO2 fixation in plants, algae, and photosynthetic bacteria. Despite of the important function in the global carbon cycle, RuBisCO suffers from a slow reaction rate and a competing reaction with O2 which draw attentions to improve the enzyme efficiency. In this study, a RuBisCO dimer from Rhodospirillum rubrum was assembled on a DNA scaffold using a dimeric DNA binding protein as an adaptor. The enzyme assembly was characterized by atomic force microscopy and RuBisCO assembled on the DNA scaffold showed avid enzymatic activity with retaining its parent carboxylase function. To mimic the environment of the natural microcompartment in cyanobacterial carboxysome that encapsulate the second enzyme carbonic anhydrase (CA) with RuBisCO, RuBisCO was next co-assembled with CA on the DNA scaffold. Although the natural carboxysome assembly is believed to enhance the RuBisCO activity, the co-assembly of RuBisCO and CA reduced the RuBisCO activity, suggesting that the preferential CO2 dehydration by CA reduced the RuBisCO reaction rate. In line with the recent study, our results suggest that the proximity in the interenzyme distance of RuBisCO and CA is not the crucial determinant for the enhanced RuBisCO activity in carboxysome. The assembly of RuBisCO and CA on DNA scaffold provides a platform for further study on the spatial control of RuBisCO and associating enzymes. 相似文献
3.
Detached wheat leaves were illuminated in air until a steady rate of photosynthesis was established. Then the gas was changed to 1% O2, 99% N2 and after 2.5 h further illumination the capacity of the leaves for photosynthesis in air was decreased to approximately 50%. Measurement of RuBP carboxylase activity in extracts showed that inhibition of photosynthesis was accompanied by 70% inactivation of this enzyme. The capacity for photosynthesis and the activity of RuBP carboxylase were recovered when leaves were returned to normal air. Extracts of the leaves made when photosynthesis and carboxylase activity were low, recovered most of the lost carboxylase activity when supplemented with bicarbonate and magnesium ions. The time courses for activation and inactivation of the RuBP carboxylase in these experiments suggests the operation of a mechanism that has not yet been elucidated. 相似文献
4.
In light and in darkness, exposure of leaf segments to CO2-free atmospheres caused a marked reduction in extractable RuBP carboxylase activity. By contrast, darkness caused a relatively small decrease in carboxylase activity in extracts from leaf segments kept in air containing CO2. Recovery of carboxylase activity in leaves during illumination in air after exposure to CO2-free conditions paralleled recovery of capacity for photosynthesis; in darkness recovery of carboxylase activity in leaves was slower than in the light. Extracts from leaves exposed to CO2-free conditions recovered activity when provided with CO2 and Mg2+; there were clearly, however, substances in the extracts that modified the activity achieved and caused anomalous decreases and increases with time after extraction. Studies of the effect of orthophosphate on the activity of purified wheat carboxylase in vitro were consistent with the view that many of the effects observed on the activity of crude leaf extracts were due to orthophosphate content. 相似文献
5.
Extracts from sunflower leaves possess a high ribulose-1,5-bisphosphate (RuBP) carboxylase capacity but this enzyme activity is not stable. A purification procedure, developed with preservation of carboxylase activity by MgSO4, yielded purified RuBP carboxylase with high specific activity (40 nkat mg-1 protein). Measurement of kinetic parameters showed high Km values (RuBP, HCO
3
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) and high Vmax of the reaction catalyzed by this sunflower enzyme; the results are compared with those obtained for soybean carboxylase. Enzyme characteristics are discussed in relation to stabilization and activation procedures and to the high photosynthesis rates of this C3 species. 相似文献
6.
Reversible light-activation of ribulose bisphosphate carboxylase/oxygenase in isolated barley protoplasts and chloroplasts 总被引:1,自引:2,他引:1
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Sicher RC 《Plant physiology》1982,70(2):366-369
The enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase displayed near-maximal activity in isolated, intact barley (Hordeum vulgare L. cv. Pennrad) mesophyll protoplasts. The carboxylase deactivated 40 to 50% in situ when protoplasts were dark-incubated 20 minutes in air-equilibrated solutions. Enzyme activity was fully restored after 1 to 2 minutes of light. Addition of 5 millimolar NaHCO3 to the incubation medium prevented dark-inactivation of the carboxylase. There was no permanent CO2-dependent activation of the protoplast carboxylase either in light or dark. Activation of the carboxylase from ruptured protoplasts was not increased significantly by in vitro preincubation with CO2 and Mg2+. In contrast to the enzyme in protoplasts, the carboxylase in intact barley chloroplasts was not fully reactivated by light at atmospheric CO2 levels. The lag phase in carbon assimilation was not lengthened by dark-adapting protoplasts to low CO2 demonstrating that light-activation of the carboxylase was not involved in photosynthetic induction. Irradiance response curves for reactivation of the the carboxylase and for CO2 fixation by isolated barley protoplasts were similar. The above results show that there was a fully reversible light-activation of the carboxylase in isolated barley protoplasts at physiologically significant CO2 levels. 相似文献
7.
The carboxylase activity of Rubisco and the photosynthetic performance in aquatic plants 总被引:3,自引:0,他引:3
Summary Activated carboxylase activities of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), as well as photosynthetic rates were measured for 42 species of freshwater and marine macrophytes. While the carboxylase activity varied greatly among the species investigated (0.2–12.5 mol CO2 mg–1 chlorophyll min–1), the submersed freshwater plants showed significantly lower activities than emergent, floating leaved or secondary submersed forms. The variability in photosynthetic rates correlated with the carboxylase activity only for the marine macroalgae, and their photosynthesis to carboxylase activity ratios were close to 1. These plants also had a consistently high inorganic carbon transport capability, and it is suggested that ribulose-1,5-bisphosphate carboxylase/oxygenase activity is an important internal factor regulating the photosynthetic capacity within this plant group where, apparently, the internal CO2 concentration is high and photorespiration is suppressed. Among the freshwater forms, it appears that their much lower inorganic carbon transport ability, rather than their carboxylase activity, limits the photosynthetic process. 相似文献
8.
1. Cell-free extracts of the photosynthetic bacterium Chlorobium thiosulfatophilum, strains 8327 and Tassajara, were assayed for ribulose 1,5-diphosphate (RuDP) carboxylase and phosphoribulokinase-the two enzymes peculiar to the reductive pentose phosphate cycle. 2. RuDP carboxylase was consistently absent in strain 8327. The Tassajara strain showed a low RuDP-dependent CO2 fixation activity that was somewhat higher in cells following transatlantic air shipment than in freshly grown cells. The stability and behaviour of this activity in sucrose density gradients were similar to those described by other workers. 3. The radioactive carboxylation products formed in the presence of RuDP by enzyme preparations from the Tassajara strain did not include 3-phosphoglycerate-the known product of the RuDP carboxylase reaction, but instead consisted of the unrelated acids glutamate, aspartate and malate. 4. Phosphoribulokinase was absent in all preparations of the two Chlorobium strains tested. By contrast, phosphoribulokinase as well as RuDP carboxylase were readily demonstrated in preparations from pea chloroplasts and the photosynthetic bacterium Rhodospirillum rubrum. 5. It is concluded that C. thiosulfatophilum appears to lack RuDP carboxylase, phosphoribulokinase, and hence, the reductive pentose phosphate cycle.Support of a J. S. Guggenheim Fellowship is gratefully acknowledged 相似文献
9.
Ribulose diphosphate oxygenase. V. Presence in ribulose diphosphate carboxylase from Rhodospirillum rubrum 总被引:2,自引:0,他引:2
F J Ryan S O Jolly N E Tolbert 《Biochemical and biophysical research communications》1974,59(4):1233-1241
Ribulose-1,5-diphosphate carboxylase was purified fifteenfold from Rhodospirillum rubrum grown autotrophically under H2 and CO2. There was RuDP oxygenase activity associated with the carboxylase. The oxygenase had maximal activity at pH 9.4. Although these bacterial RuDP oxygenase and carboxylase activities were cold labile, activity could not be restored by treatment at 50° in the presence of Mg++ and a sulfhydryl reagent, in contrast to results with the enzyme from eukaryotes. 相似文献
10.
Ribulose 1,5-bisphosphate and activation of the carboxylase in the chloroplast 总被引:11,自引:9,他引:2
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Ribulose 1,5-bisphosphate in the chloroplast has been suggested to regulate the activity of the ribulose bisphosphate carboxylase/oxygenase. To generate high levels of ribulose bisphosphate, isolated and intact spinach chloroplasts were illuminated in the absence of CO2. Under these conditions, chloroplasts generate internally up to 300 nanomoles ribulose 1,5-bisphosphate per milligram chlorophyll if O2 is also absent. This is equivalent to 12 millimolar ribulose bisphosphate, while the enzyme, ribulose bisphosphate carboxylase, offers up to 3.0 millimolar binding sites for the bisphosphate in the chloroplast stroma. During illumination, the ribulose bisphosphate carboxylase is deactivated, due mostly to the absence of CO2 required for activation. The rate of deactivation of the ribulose bisphosphate carboxylase was not affected by the chloroplast ribulose bisphosphate levels. Upon addition of CO2, the carboxylase in the chloroplast was completely reactivated. Of interest, addition of 3-phosphoglycerate stopped deactivation of the carboxylase in the chloroplast while ribulose bisphosphate accumulated. With intact chloroplasts in light, no correlation between deactivation of the carboxylase and ribulose bisphosphate levels could be shown. 相似文献
11.
The catalytic subunit of cyclic AMP-dependent protein kinase stimulates the inactivation of acetyl-coenzyme A (CoA) carboxylase by acetyl-CoA carboxylase kinase. The stimulated inactivation of carboxylase is due to activation of carboxylase kinase by the catalytic subunit. Activation of carboxylase kinase activity is accompanied by the incorporation of 0.6 mol of phosphate per mole of carboxylase kinase. Addition of the regulatory subunit of cyclic AMP-dependent protein kinase prevents the activation of carboxylase kinase. Phosphorylation and activation of carboxylase kinase has no effect on the Km for ATP, but decreases the Km for acetyl-CoA carboxylase from 93 to 45 nm. Inactivation of carboxylase by the carboxylase kinase requires the presence of coenzyme A even when the activated carboxylase kinase is used. Acetyl-CoA carboxylase is not phosphorylated or inactivated by the catalytic subunit of cyclic AMP-dependent protein kinase. 相似文献
12.
Ribulose 1,5-bisphosphate carboxylase/oxygenase purified from malate-grown Thiocapsa roseopersicina required Mg2+ for the activation of both carboxylase and oxygenase activities. Mg2+ was either not required or required at very low concentrations for catalysis by both enzyme activities. EDTA and dithiothreitol had no effect on ribulose 1,5-biphosphate oxygenase. The K0.5 values with respect to Mg2+ for activation of the carboxylase and oxygenase activities were 8.4 and 2 mm, respectively. Ribulose 1,5-biphosphate carboxylase and oxygenase activities revealed differential sensitivities to 6-phosphogluconate. This ligand at 1 mm inhibited the carboxylase activity 30%, whereas the oxygenase activity was inhibited by 69%. 相似文献
13.
Properties of phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31) obtained from isolated guard-cell protoplasts of Vicia faba L. were determined following rapidly desalting of the extract on a Sephadex G 25 column. The activity of PEP carboxylase was measured as a function of PEP and malate concentration, pH and K+ concentration within 2–3 min after homogenization of the guard-cell protoplasts. The activity of this enzyme was stimulated by PEP concentrations of 0.1 to 0.75 mM and by K+ ions (12 mM), but inhibited by PEP concentrations above 1 mM and by malate. Changes in the Km(PEP) and Vmax values with increasing malate concentrations (2.5 and 5 mM) indicate that the malate level, varying in relation to the physiological state of guard cells, plays an important role in regulating the properties of phosphoenolpyruvate carboxylase.Abbreviations CAM
Crassulacean acid metabolism
- GCP
guard-cell protoplast
- PEP
phosphoenolpyruvate
Dedicated to Professor Dr. Hubert Ziegler on the occasion of his 60th birthday 相似文献
14.
Heterotrophically grown Euglena synthesize grains of paramylon, its reserve carbohydrate, in a vesicular complex of mitochondrial origin. A CO2 fixation activity in dark grown Euglena was demonstrated in the mitochondria via paramylon. At the beginning of the exponential phase of growth, the activity of phosphoenolpyruvate carboxykinase increases before the augmentation of paramylon.At the end of the exponential phase, the activity of this enzyme decreases, and low residual levels persist in the transition and stationary phases of growth. The activity of phosphoenolpyruvate carboxylase evolves inversely during the heterotrophic growth of the algae in succinate- or a lactate-containing medium. A compartmentalized scheme of carbon metabolism in mitochondria is presented.Abbreviations PEP
phosphoenolpyruvate
- OAA
oxaloacetate
- PGA
phosphoglyceric acid 相似文献
15.
C. K. M. Rathnam 《Planta》1978,141(3):289-295
The activity of phosphoenolpyruvate (PEP) carboxylase EC 4.1.1.31 in leaf extracts of Eleusine indica L. Gaertn., a C4 plant, exhibited a temperature optimum of 35–37° C with a complete loss of activity at 50° C. However, the enzyme was protected effectively from heat inactivation up to 55° C by L-aspartate. Activation energies (Ea) for the enzyme in the presence of aspartate were 2.5 times lower than that of the control enzyme. Arrhenius plots of PEP carboxylase activity (±aspartate) showed a break in the slope around 17–20° C with a 3-fold increase in the Ea below the break. The discontinuity in the slopes was abolished by treating the enzyme extracts with Triton X-100, suggesting that PEP carboxylase in C4 plants is associated with lipid and may be a membrane bound enzyme. Depending upon the species, the major C4 acid formed during photosynthesis (malate or aspartate) was found to be more protective than the minor C4 acid against the heat inactivation of their PEP carboxylase. Oxaloacetate, the reaction product, was less effective compared to malate or aspartate. Several allosteric inhibitors of PEP carboxylase were found to be moderately to highly effective in protecting the C4 enzyme while its activators showed no significant effect. PEP carboxylase from C3 species was not protected from thermal inactivation by the C4 acids. The physiological significance of these results is discussed in relation to the high temperature tolerance of C4 plants.Abbreviations CAM
crassulaccan acid metabolism
- Chl
chlorophyll
- Ea
activation energy
- PEP
phosphoenolypyruvate
Journal Series Paper, New Jersey Agricultural Experiment Station 相似文献
16.
Biosynthesis of ribulose-1,5-bisphosphate carboxylase in spinach leaf protoplasts 总被引:2,自引:1,他引:1
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Spinach leaf (Spinacia oleracea L. var. Kyoho) protoplasts sustain protein-synthesizing activity as measured by the incorporation of [14C]-leucine into the protein fraction both in the light and in the dark. By the immunoprecipitation of ribulose-1,5-bisphosphate (RuP2) carboxylase with rabbit antibody raised against the purified spinach enzyme preparation, it was found that approximately 7% of the total radiocarbon incorporated into the protein fraction in the light was in the carboxylase molecules. However, there was no measurable net increase observed in the content of the enzyme protein in the experimental conditions employed. It was found that both chloramphenicol and cycloheximide inhibited the incorporation of [14C]leucine into RuP2 carboxylase and its constituent subunits, as measured by the immunoprecipitation of the enzyme molecule and its subunits, A and B. 相似文献
17.
Sanjay K. Gupta Maurice S. B. Ku Jenq-Horng Lin Dianzhong Zhang Gerald E. Edwards 《Photosynthesis research》1994,42(2):133-143
In this report, the effects of light on the activity and allosteric properties of phosphoenolpyruvate (PEP) carboxylase were examined in newly matured leaves of several C3 and C4 species. Illumination of previously darkened leaves increased the enzyme activity 1.1 to 1.3 fold in C3 species and 1.4 to 2.3 fold in C4 species, when assayed under suboptimal conditions (pH 7) without allosteric effectors. The sensitivities of PEP carboxylase to the allosteric effectors malate and glucose-6-phosphate were markedly different between C3 and C4 species. In the presence of 5 mM malate, the activity of the enzyme extracted from illuminated leaves was 3 to 10 fold higher than that from darkened leaves in C4 species due to reduced malate inhibition of the enzyme from illuminated leaves, whereas it increased only slightly in C3 species. The Ki(malate) for the enzyme increased about 3 fold by illumination in C4 species, but increased only slightly in C3 species. Also, the addition of the positive effector glucose-6-phosphate provided much greater protection against malate inhibition of the enzyme from C4 species than C3 species. Feeding nitrate to excised leaves of nitrogen deficient plants enhanced the degree of light activation of PEP carboxylase in the C4 species maize, but had little or no effect in the C3 species wheat. These results suggest that post-translational modification by light affects the activity and allosteric properties of PEP carboxylase to a much greater extend in C4 than in C3 species. 相似文献
18.
Pooja K Strope Kenneth W Nickerson Steven D Harris Etsuko N Moriyama 《BMC evolutionary biology》2011,11(1):80
Background
Urea amidolyase breaks down urea into ammonia and carbon dioxide in a two-step process, while another enzyme, urease, does this in a one step-process. Urea amidolyase has been found only in some fungal species among eukaryotes. It contains two major domains: the amidase and urea carboxylase domains. A shorter form of urea amidolyase is known as urea carboxylase and has no amidase domain. Eukaryotic urea carboxylase has been found only in several fungal species and green algae. In order to elucidate the evolutionary origin of urea amidolyase and urea carboxylase, we studied the distribution of urea amidolyase, urea carboxylase, as well as other proteins including urease, across kingdoms. 相似文献19.
Acetyl-coenzyme A carboxylase in maize leaves 总被引:6,自引:0,他引:6
Purified chloroplasts from mesophyll and bundle sheath cells of maize leaves have been shown to be the location of acetyl-CoA carboxylase. In disrupted chloroplasts the enzyme was recovered in the stromal fraction, along with protein-bound biotin; acetyl-CoA carboxylase activity did not require a membrane component. Mg2+ and ATP are required for activity and sulfhydryl protecting agents enhance stability of the enzyme. Acetyl-CoA carboxylase activity was independent of leaf development in cell-free extracts of maize. Comparison of acetyl-CoA carboxylase activity with [14C]acetate incorporation into lipids, in isolated chloroplasts from developing leaves of maize, indicate that acetyl-CoA carboxylase is not limiting fatty acid synthesis. 相似文献
20.
Characterization of biotin and 3-methylcrotonyl-coenzyme a carboxylase in higher plant mitochondria 总被引:4,自引:3,他引:1
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Mitochondria from green pea (Pisum sativum) leaves were purified free of peroxisomes and chlorophyll contamination and examined for their biotin content. The bulk of the bound biotin detected in plant mitochondria was shown to be associated with the matrix space to a concentration of about 13 micromolar, and no free biotin was detected. Western blot analysis of mitochondrial polypeptides using horseradish peroxidase-labeled streptavidin revealed a unique biotin-containing polypeptide with a molecular weight of 76,000. This polypeptide was implicated as being the biotinylated subunit of 3-methylcrotonyl-coenzyme A (CoA) carboxylase. Fractionation of pea leaf protoplasts demonstrated that this enzyme activity was located largely in mitochondria. The 3-methylcrotonyl-CoA carboxylase activity was latent when assayed in isotonic media. The majority of the enzyme activity was found in the soluble matrix of mitochondria. Maximal 3-methylcrotonyl-CoA carboxylase activity was found at pH 8.3 in the presence of Mg2+. Kinetic constants (apparent Km values) for the enzyme substrates were: 3-methylcrotonyl-CoA, 0.05 millimolar; ATP, 0.16 millimolar; HCO3−, 2.2 millimolar. The involvement of 3-methylcrotonyl-CoA carboxylase in the leucine degradation pathway in plant mitochondria is proposed. 相似文献