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1.
Sugarcane (Saccharum officinarum L.) leaf parenchyma cells bathed in 1X solution maintained an average membrane potential of −135 millivolts in the dark. No difference in membrane potential was found between clones 51 NG 97 and H50 7209. An electrogenic pump appears to contribute to membrane potential in these cells. Sugars (25 millimolar) added externally caused the following membrane potential depolarizations (in millivolts) in clone 51 NG 97: glucose, 18 ± 4; galactose, 24 ± 7; 3-O-methylglucose, 10 ± 4; sucrose, 22 ± 3; fructose, 21 ± 7; raffinose, 9 ± 3; mannitol, 0; lactose, 0; melibiose, 0; and 1-O-methyl-α-galactose, 0. Glycine (25 millimolar) and serine (10 millimolar) caused depolarizations of 47 ± 7 and 23 ± 2 millivolts, respectively. Depolarization shows saturation kinetics with respect to glucose concentration, with a Km of 3 to 6 millimolar. The metabolic inhibitors KCN and salicyl hydroxamic acid together caused depolarization of the membrane potential and greatly inhibited depolarization by 25 millimolar glucose and 25 millimolar raffinose. In a series of substitution experiments, glucose (25 millimolar) caused almost total inhibition of depolarization by raffinose, sucrose, and 3-O-methylglucose (all 25 millimolar), but only partial inhibition of depolarization to 25 millimolar glycine. Glycine (25 millimolar), also, only partially inhibited depolarization by 25 millimolar glucose. Total depolarization to 25 millimolar glycine and 25 millimolar glucose was comparable to the amount of depolarization of membrane potential caused by 1 millimolar KCN plus 1 millimolar salicyl hydroxamic acid. The results are consistent with a co-transport mechanism of membrane transport, with sugars and amino acids being transported by separate carrier systems.  相似文献   

2.
Van Den Bossche H. and De Nollin S. 1973. Effects of mebendazole on the absorption of low molecular weight nutrients by Ascaris suum. International Journal for Parasitology3: 401–407. The effect of the anthelmintic drug, mebendazole, on the uptake and/or transport of glucose, fructose, 3-O-methylglucose, glycine, proline, methionine and palmitic acid was studied on in vitro incubated Ascaris suum. The experiments presented indicate that mebendazole inhibits the uptake and/or transport of glucose by A. suum. This inhibition is followed by a marked decrease in the glycogen content of the ascaris muscle. The addition of glucose to the incubation medium significantly enhanced the rate of uptake and/or transport of 3-O-methylglueose, glycine, methionine, proline and palmitic acid indicating that the absorption mechanisms depend on energy.Therefore, the inhibitory effect of mebendazole on the glucose uptake also results in a decreased uptake of 3-O-methylglucose and of the amino acids and fatty acid studied. The fructose uptake was not affected by the addition of glucose.Although mebendazole decreased the uptake of the hexoses and of the amino acids whether or not glucose was added, the uptake of palmitic acid was not affected when glucose was omitted from the medium. Mebendazole failed to exhibit an effect on the uptake, transport and/or utilization of glucose in rat.  相似文献   

3.
Maynard JW  Lucas WJ 《Plant physiology》1982,70(5):1436-1443
Concentration curves for sugar and amino acid uptake by Beta vulgaris L. leaf tissues contained both a saturable and a linear component. Similarly shaped curves were obtained for influx of sucrose, glucose, and 3-O-methyl glucose by leaf discs, whole petiole slices, petiole segments containing pith tissue only, and petiole segments containing vascular bundles, although the tissues took up the various sugars via different proportions of saturable versus linear uptake. Two millimolar p-chloromercuribenzenesulfonic acid selectively inhibited the saturable component of sucrose uptake, but had almost no effect on the linear component. Uptake of glucose and 3-O-methyl glucose remained unaffected by p-chloromercuribenzenesulfonic acid treatment. Anoxia was found to inhibit the linear component of both sucrose and 3-O-methyl glucose influx, while the saturable component remained unaffected. The linear component of sucrose uptake was also competitively inhibited by maltose, as well as being selectively promoted by certain exposures to 5 millimolar N-ethylmaleimide, 2 micrograms per milliliter cycloheximide, and high levels of mannitol acting as osmoticum. These results support the proposal that the linear component is due to a process more complex than simple, or exchange, diffusion. It would also appear that the linear transport component utilizes a separate energy source than does the saturable component of sucrose influx.

Evidence for phloem loading from the apoplast was re-examined with respect to the present findings. Saturable sucrose uptake by minor vein tissues may represent retrieval of solute from the free space, which could explain the `apoplastic loading' phenomenon.

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4.
In vitro sugar transport into developing isolated maize embryos was studied. Embryo fresh and dry weight increased concomitantly with endogenous sucrose concentration and glucose uptake throughout development. However, endogenous glucose and fructose concentration and sucrose uptake remained constant. The uptake kinetics of radiolabeled sucrose, glucose, and fructose showed a biphasic dependence on exogenous substrate concentration. Hexose uptake was four to six times greater than sucrose uptake throughout development. Carbonylcyanide-m-chlorophenylhydrazone and dinitrophenol inhibited sucrose and glucose uptake significantly, but 3-O-methyl glucose uptake was less affected. The uptake of 1 millimolar sucrose was strongly pH dependent while glucose was not. Glucose and fructose were readily converted to sucrose and insoluble products soon after absorption into the embryo. Thus, sucrose accumulated, while glucose pools remained low. Based on the findings of this and other studies a model for sugar transport in the developing maize kernel is presented.  相似文献   

5.
Evidence for a plasmalemma redox system in sugarcane   总被引:1,自引:1,他引:0       下载免费PDF全文
A plasmalemma-bound NADH-dependent redox system has been identified in protoplasts isolated from cell suspensions of sugarcane. This system oxidized NADH as well as NADPH, increased O2 consumption 3-fold, and increased the pH of the external medium while the cytoplasmic pH was decreased. In the presence of NADH, ferricyanide was rapidly reduced and the external medium was acidified. The uptake rates of K+, 3-O-methylglucose, leucine, and arginine were all decreased in the presence of NADH.  相似文献   

6.
Suspensions of dark-adapted guard cell protoplasts of Vicia faba L. alkalinized their medium in response to irradiation with red light. The alkalinization peaked within about 50 minutes and reached steady state shortly thereafter. Simultaneous measurements of O2 concentrations and medium pH showed that oxygen evolved in parallel with the red light-induced alkalinization. When the protoplasts were returned to darkness, they acidified their medium and consumed oxygen. Both oxygen evolution and medium alkalinization were inhibited by 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU). In photosynthetically competent preparations, light-dependent medium alkalinization is diagnostic for photosynthetic carbon fixation, indicating that guard cell chloroplasts have that capacity. The striking contrast between the responses of guard cell protoplasts to red light, which induces alkalinization, and that to blue light, which activates proton extrusion, suggests that proton pumping and photosynthesis in guard cells are regulated by light quality.  相似文献   

7.
The effects of ATP on glucose transport and metabolism were studied in rat adipocytes. Over a concentration range of 10–250 μm, ATP was found to inhibit several aspects of adipocyte glucose metabolism, particularly when stimulated by insulin. Much of the effect of ATP on glucose metabolism appeared related to impairment of glucose transport, reflected by inhibition of both basal and insulin-stimulated rates of 3-O-methylglucose transport. ATP inhibited the V of insulin-stimulated 3-O-methylglucose transport, but had no effect on the Km. The inhibitory effects of ATP were much less apparent when cells were preincubated with insulin, suggesting that ATP inhibited only the components of hexose transport not yet activated by the hormone. At very high medium glucose concentrations, where transport was no longer rate limiting for metabolism, there was no inhibition of glucose oxidation by 250 μm ATP. However, when hexose transport was blocked with cytochalasin B (50 μm), a small inhibitory effect of ATP persisted on basal and insulin-stimulated glucose and fructose oxidation, suggesting that intracellular metabolism was impaired. The mechanism of the intracellular effect did not appear to be caused by uptake of exogenous ATP. These studies provide further evidence that energy metabolism may play an important role in the regulation of facilitated glucose transport.  相似文献   

8.
Daie J 《Plant physiology》1987,84(4):1033-1037
Phloem tissue isolated from celery (Apium graveolens L.) was used to investigate the regulation of sucrose uptake by turgor (manipulated by 50-400 milliosomolal solutions of polyethylene glycol) and hormones indoleacetic acid (IAA) and gibberillic acid (GA3). Sucrose uptake was enhanced under low cellular turgor (increase in the Vmax). Furthermore, enhancement of sucrose uptake was due to a net increase in influx rates since sucrose efflux was not affected by cell turgor. Manipulations of cell turgor had no effect on 3-O-methyl glucose uptake. When 20 millimolar buffer was present in uptake solutions, low turgor-induced effects were observed only at low pH range (4.5-5.5). However, the effect was extended to higher external pH (up to 7.5) when buffer was omitted from uptake solutions. A novel interaction between cellular turgor and hormone treatments was observed, in that GA3 (10 micromolar) and IAA (0.1-100 micromolar) enhanced sucrose uptake only at moderate turgor levels. The hormones elicited little or no response on sucrose uptake under conditions of low or high cell turgor. Low cell turgor, IAA, GA3, and fusicoccin caused acidification by isolated phloem segments in a buffer-free solution. It is suggested that enhanced sucrose uptake in response to low turgor and/or hormones was mediated through the plasmalemma H+-ATPase and most likely occurred at the site of loading.  相似文献   

9.
Evidence for a specific glutamate/h cotransport in isolated mesophyll cells   总被引:1,自引:1,他引:0  
Mechanically isolated Asparagus sprengeri Regel mesophyll cells were suspended in 1 millimolar CaSO4. Immediate alkalinization of the medium occured on the addition of 1 millimolar concentrations of l-glutamate (Glu) and its analog l-methionine-d,l-sulfoximine (l-MSO). d-Glu and the l isomers of the protein amino acids did not elicit alkalinization. l-Glu dependent alkalinization was transient and acidification resumed after approximately 30 to 45 minutes. At pH 6.0, 5 millimolar l-Glu stimulated initial rates of alkalinization that varied between 1.3 to 4.1 nmol H+/106 cells·minute. l-Glu dependent alkalinization was saturable, increased with decreasing pH, was inhibited by carbonyl cyanide-p-trichloromethoxyphenyl hydrazone (CCCP), and was not stimulated by light. Uptake of l-[U-14C]glutamate increased as the pH decreased from 6.5 to 5.5, and was inhibited by l-MSO. l-Glu had no influence on K+ efflux. Although evidence for multiple amino acid/proton cotransport systems has been found in other tissues, the present report indicates that a highly specific l-Glu/proton uptake process is present in Asparagus mesophyll cells.  相似文献   

10.
Energetics of Amino Acid Uptake by Vicia faba Leaf Tissues   总被引:7,自引:5,他引:2  
The uptake of [U-14C]threonine and of (α-14C]aminoisobutyrate (α-AIB) by Vicia faba leaf discs is strongly pH dependent (optimum: pH 4.0) and exhibits biphasic saturation kinetics. Kinetics of α-AIB uptake at different pH values indicate that acidic pH values decrease the Km of the carriers while the maximal velocity remains nearly unaffected. Similar results were obtained for both system 1 (from 0.5 to 5 millimolar) and system 2 (from 20 to 100 millimolar).

After addition of amino acids to a medium containing leaf fragments, alkalinizations depending both on the amino acid added and on its concentration have been recorded.

The effects of compounds which increase (fusicoccin) or decrease (uncouplers, ATPase inhibitors, high KCl concentrations) the protonmotive force were studied both on the acidification of the medium and on amino acid uptake by the tissues. There is a close relationship between the time required for the effect of these compounds on the acidification and that needed for inhibition of uptake.

Studies with thiol inhibitors show that 0.1 millimolar N-ethylmaleimide preferentially inhibits uptake by the mesophyll whereas 0.1 millimolar parachloromercuribenzenesulfonate affects rather uptake by the veins.

New evidence was found which added to the electrophysiological data already supporting the occurrence of proton amino acid symport in leaf tissues, particularly in the veins.

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11.
Vicia faba leaf discs without epidermis were pretreated with parachloromercuribenzenesulfonic acid (PCMBS), rinsed and incubated on [14C]sucrose (1 or 40 millimolar). Those sucrose concentrations were chosen as representative of the apparent uptake system 1 (1 millimolar) and system 2 (40 millimolar) previously characterized. Pretreatment with 0.5 millimolar PCMBS for 20 minutes inhibited system 1 and system 2 by about 70%.

Addition of unlabeled sucrose during PCMBS-pretreatment protected the carrier(s) from the inhibition, whereas glucose, fructose, and sucrose analogs were unable to afford protection. At 1 millimolar [14C]sucrose, the protection resulted in a small but consistent reduction of normal inhibition (from 63 to 45%) for sucrose concentrations of 50 millimolar and more during pretreatment. Contrarily, at 40 millimolar [14C]sucrose, the protection increased linearly with the sucrose concentration in the pretreatment medium, and complete prevention of inhibition was reached for 250 millimolar sucrose.

The protection was not due to exchange diffusion and was located in the veins. Michaelian kinetics indicated that PCMBS and sucrose compete with each other at the active site of the carrier.

Among 14 compounds tested (sugars, amino-acids, hormones, 32P), sucrose uptake was by far the most sensitive to PCMBS. Sucrose preferentially protected its carrier(s) from inhibition. Treatment with 20 millimolar cysteine or 20 millimolar dithioerythreitol reversed inhibition by PCMBS pretreatment.

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12.
Cardiac myocytes were isolated from adult rat ventricles by a method which preserves their functional integrity, including long survival in physiological concentrations of Ca2+. Sarcolemmal glucose transport was assessed by measuring linear initial uptake rates of the nonmetabolized glucose analog3-O-methyl-d-glucose. Transport was saturable and showed competition byd-glucose and other features of chemical and stereo-selectivity. Transport was stimulated by insulin in a dose-dependent manner, resulting in an almost 5-fold increase inVmax, with little change inKm. Stimulation of 3-methylglucose transport by insulin was largely Ca2+ -dependent. Omission of Ca2+ from the incubation medium caused a minor rise in basal 3-methylglucose uptake but the insulin-stimulated rise inVmax was only 30%. The Ca2+ antagonist D600 also antagonized stimulation of hexose transport by insulin. In all the above respects, 3-methylglucose transport in myocytes is identical to that in intact heart muscle. In addition, the decrease in insulin response by Ca2+ emission was partially reversed by subsequent return to a Ca2+ -containing medium. ATP levels remained stable in the absence of Ca2+, showing that the Ca2+ dependence did not reflect nonspecific cell damage.  相似文献   

13.
Sugar transport in isolated corn root protoplasts   总被引:7,自引:6,他引:1       下载免费PDF全文
Isolated corn (Zea mays L.) root protoplasts were used to study sucrose and hexose uptake. It is found that glucose was preferentially taken up by the protoplasts over sucrose and other hexoses. Glucose uptake showed a biphasic dependence on external glucose concentration with saturable (Km of 7 millimolar) and linear components. In contrast, sucrose uptake only showed a linear kinetic curve. Sucrose and glucose uptake were linear over a minimum of 1 hour at pH 6.0 and 1 millimolar exogenous sugar concentration. Glucose uptake showed a sharp 42°C temperature optimum, while sucrose uptake showed a lower temperature sensitivity which did not reach a maximum below 50°C. Uptake of both sugars was sensitive to several metabolic inhibitors and external pH. Differences between sucrose and glucose uptake in two different sink tissue (i.e. protoplasts from corn roots and soybean cotyledons) are discussed.  相似文献   

14.
Addition of l-[U-14C]glutamate to a suspension of mechanically isolated asparagus (Asparagus sprengeri Regel) mesophyll cells results in (a) alkalinization of the medium, (b) uptake of l-[U-14C]glutamate, and (c) efflux of [14C]4-aminobutyrate, a product of glutamate decarboxylation. All three phenomena were eliminated by treatment with 1 millimolar aminooxyacetate. In vitro glutamate decarboxylase (GAD) assays showed that (a) 2 millimolar aminooxyacetate eliminated enzyme activity, (b) activity was pyridoxal phosphate-dependent, and (c) activity exhibited a sharp pH optimum at 6.0 that decreased to 20% of optimal activity at pH 5.0 and 7.0. Addition of 1.5 millimolar sodium butyrate or sodium acetate to cell suspensions caused immediate alkalinization of the medium followed by a resumption of acidification of the medium at a rate approximately double the initial rate. The data indicate that (a) continued H+/l-glutamate contransport is dependent upon GAD activity, (b) the pH-dependent properties of GAD are consistent with a role in a metabolic pH-stat, and (c) the regulation of intracellular pH during H+/l-Glu symport may involve both H+ consumption during 4-aminobutyrate production and ATP-driven H+ efflux.  相似文献   

15.
A Technique for Collection of Exudate from Pea Seedlings   总被引:2,自引:2,他引:0  
Ethylenediaminetetraacetic acid (EDTA), at concentrations higher than 1.0 millimolar, is phytotoxic to etiolated seedlings of Pisum sativum. Substantial vascular exudation from pea epicotyls could be obtained without tissue damage at 0.5 millimolar EDTA if the solution was buffered at pH 7.5 with sodium N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid. Treated seedlings exuded 950 micrograms (leucine equivalents) of ninhydrin-positive material per day and 870 micrograms (glucose equivalents) of anthrone-positive material per day. Amino acid analysis showed the exudate to have glutamine as the major amido nitrogen containing compound and sucrose was shown to be the major sugar. Radiolabeled tryptophan and sucrose applied to cotyledons were transferred through the epicotyl and into the collection medium. The pH profile for exudation shows half maximal exudation at pH 7.2, indicating the promotion of exudation by EDTA is probably not due simply to Ca2+ chelation.  相似文献   

16.
Replacement of mannitol with sucrose decreases the binding of [203Hg]-p-chloromercuribenzenesulphonic acid (PCMBS) to Vicia faba leaf discs without epidermis. This decrease is optimal for 20 minutes on incubation, is concentration-dependent, and is also found with maltose and raffinose. In parallel experiments, the addition of sucrose, maltose, and raffinose during PCMBS pretreatment was shown to increase subsequent uptake of [U-14C]sucrose. In contrast, d- or l-glucose, 3-O-methylglucose, galactose, fructose, palatinose, turanose, or melibiose had no effect either on PCMBS binding or on [14C]sucrose uptake. The sucrose-induced decrease of PCMBS binding is retained after a cold and ionic shock. Measurements of specific activities of membrane fractions prepared from tissues incubated in labeled PCMBS show that the decrease concerns the 120,000 gravity pellet, but that very mild procedures must be chosen to prevent redistribution of label in the supernatant. Altogether, the data provide new support to the hypothesis that the active site of the sucrose carrier contains a group sensitive to PCMBS.  相似文献   

17.
To evaluate and characterize stevioside biosynthetic pathway in Stevia rebaudiana Bertoni cv Houten, two enzyme fractions that catalyze glucosylation of steviol (ent-13-hydroxy kaur-16-en-19-oic acid) and steviol-glucosides (steviol-13-O-glucopyranoside, steviolbioside and stevioside), utilizing UDP-glucose as the glucose donor, were prepared from the soluble extracts of S. rebaudiana leaves. Enzyme fraction I, passed through DEAE-Toyopearl equilibrated with 50 millimolar K-phosphate pH 7.5, catalyzed the glucosylation to steviol and 19-O-methylsteviol, but not to iso-steviol and 13-O-methylsteviol, indicating that 13-hydroxyl group of the steviol skeleton is glucosylated first from UDP-glucose to produce steviol-13-O-glucopyranoside. Enzyme fraction II, eluted from the DEAE-Toyopearl column with 0.15 molar KCI, catalyzed the glucose transfer from UDP-glucose to steviol-13-O-glucopyranoside, steviolbioside and stevioside, but not to rubusoside (13, 19-di-O-glucopyranoside) and rebaudioside A. The reaction products glucosylated from steviol-13-O-glucopyranoside, steviolbioside and stevioside were identified to be steviolbioside, stevioside and rebaudioside A, respectively. These results indicate that in the steviol-glucoside biosynthetic pathway, steviol-13-O-glucopyranoside produced from the steviol glucosylation is successively glucosylated to steviolbioside, then to stevioside producing rebaudioside A.  相似文献   

18.
In isolated phloem segments of celery (Apium graveolens L.), a tissue highly specific for sucrose and mannitol uptake, glucose uptake occurs at very low rates and exhibits biphasic kinetics. Nonpenetrating inhibitors such as parachloromercuribenzene sulfonic acid did not inhibit glucose uptake. However, uptake was greatly inhibited by penetrating inhibitors such as N-ethylmaleimide and carbonylcyanide-m-chlorophenyl hydrazone. Carbonylcyanide-m-chlorophenyl hydrazone inhibition of uptake was reversed by washing and addition of thiol reagents to uptake solutions. Phlorizin, a competitive inhibitor of glucose caused moderate inhibition of uptake only after 3 hours of tissue exposure. Low pH, fusicoccin, and low turgor which enhance H+-sugar cotransport did not alter uptake rates. Furthermore, glucose did not induce alkalinization of the uptake media. Efflux analysis indicated that the presence of 50 millimolar unlabeled glucose in the wash media enhanced exchange of the labeled glucose across the tonoplast. Results indicate that the glucose carrier is not located at the plasmalemma but appears to be present at the membrane of an intracellular compartment, most likely the tonoplast. Carrier-mediated glucose transport in this tissue is proposed to be a facilitated diffusion.  相似文献   

19.
Sucrose synthase of soybean nodules   总被引:6,自引:6,他引:0  
Sucrose synthase (UDPglucose: d-fructose 2-α-d-glucosyl transferase, EC 2.4.1.13) has been purified from the plant cytosolic fraction of soybean (Glycine max L. Merr cv Williams) nodules. The native enzyme had a molecular weight of 400,000. The subunit molecular weight was 90,000 and a tetrameric structure is proposed for soybean nodule sucrose synthase. Optimum activity in the sucrose cleavage and synthesis directions was at pH 6 and pH 9.5 respectively, and the enzyme displayed typical Michaelis-Menten kinetics. Soybean nodule sucrose synthase had a high affinity for UDP (Km, 5 micromolar) and a relatively low affinity for ADP (apparent Km, 0.13 millimolar) and CDP (apparent Km, 1.1 millimolar). The Km for sucrose was 31 millimolar. In the synthesis direction, UDPglucose (Km, 0.012 millimolar) was a more effective glucosyl donor than ADPglucose (Km, 1.6 millimolar) and the Km for fructose was 3.7 millimolar. Divalent cations stimulated activity in both the cleavage and synthesis directions and the enzyme was very sensitive to inhibition by heavy metals.  相似文献   

20.
The mechanism of hexose transport into plasma membrane vesicles isolated from mature sugarbeet leaves (Beta vulgaris L.) was investigated. The initial rate of glucose uptake into the vesicles was stimulated approximately fivefold by imposing a transmembrane pH gradient (ΔpH), alkaline inside, and approximately fourfold by a negative membrane potential (ΔΨ), generated as a K+-diffusion potential, negative inside. The -fold stimulation was directly related to the relative ΔpH or ΔΨ gradient imposed, which were determined by the uptake of acetate or tetraphenylphosphonium, respectively. ΔΨ- and ΔpH-dependent glucose uptake showed saturation kinetics with a Km of 286 micromolar for glucose. Other hexose molecules (e.g. 2-deoxy-d-glucose, 3-O-methyl-d-glucose, and d-mannose) were also accumulated into plasma membrane vesicles in a ΔpH-dependent manner. Inhibition constants of a number of compounds for glucose uptake were determined. Effective inhibitors of glucose uptake included: 3-O-methyl-d-glucose, 5-thio-d-glucose, d-fructose, d-galactose, and d-mannose, but not 1-O-methyl-d-glucose, d- and l-xylose, l-glucose, d-ribose, and l-sorbose. Under all conditions of proton motive force magnitude and glucose and sucrose concentration tested, there was no effect of sucrose on glucose uptake. Thus, hexose transport on the sugarbeet leaf plasma membrane was by a H+-hexose symporter, and the carrier and possibly the energy source were not shared by the plasma membrane H+-sucrose symporter.  相似文献   

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