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1.
Some methods for measuring the uptake of sugars by yeasts were investigated critically. A study was made of the effects of starvation of Pichia pinus, Candida utilis, Saccharomyces cerevisiae and Rhodosporidium toruloides on their uptake of d-glucose and 2-deoxy-d-glucose. Marked changes in the rates of uptake of these sugars occurred during 10 h of starvation, including (a) an immediate increase of up to 75% above that for growing cells and (b) a continuous decline to as little as 4%. Each yeast behaved differently. The rates did not remain constant during the periods of starvation often used for studies on the transport of sugars into yeasts. For Pichia pinus, there were striking differences, associated with starvation, between the transport of 2-deoxy-d-glucose and d-glucose, despite evidence that the two sugars enter this yeast by means of the same carrier. Some physiological explanations for these findings are discussed. 相似文献
2.
Srdjan Novak Tony D'Amore Inge Russell Graham G. Stewart 《Journal of industrial microbiology & biotechnology》1991,7(1):35-39
Summary The non-metabolizable and toxic glucose analogue 2-deoxy-d-glucose (2-DOG) has been widely employed to screen for regulatory mutants which lack catabolite repression. A number of yeast mutants resistant to 2-DOG have recently been isolated in this laboratory. One such mutant, derived from aSaccharomyces cerevisiae haploid strain, was demonstrated to be derepressed for maltose, galactose and sucrose uptake. Furthermore, kinetic analysis of glucose transport suggested that the high affinity glucose transport system was also derepressed in the mutant strain. In addition, the mutant had an increased intracellular concentration of trehalose relative to the parental strain. These results indicate that the 2-DOG resistant mutant is defective in general glucose repression. 相似文献
3.
The steady-state levels of distribution of glycine,l-aspartic acid,l-leucine and, to a lesser extent, ofl-lysine andl-methionine, in actidione-treated baker’s yeast cells are significantly altered (usually decreased) in the presence ofd-glucose,d-mannose,d-fructose, 2-deoxy-d-glucose, maltose, sucrose and, after induction,d-galactose. Stimulatory effects ofd-ribose,l-sorbose andd-xylose are not highly significant. Pronounced effects of sugars were also found anaerobically. No effect of amino acids on
sugar uptake was observed. Three types of interaction appear to be present: (1) increase of energy reserves by metabolized
sugars; (2) increased rate of carrier breakdown in the presence of metabolized sugars; (3) interaction at the carrier level
in a “heteropolyvalent” membrane complex. 相似文献
4.
The mutant R33 of the obligatory aerobic yeastRhodotorula glutinis exhibited a defect ind-glucose uptake. Detailed kinetic studies ofd-glucose andd-fructose transport in wild-type and mutant strains provided evidence for the existence in the plasma membrane of a carrier
specific for fructose. The transport ofd-fructose in the mutant exhibited saturation kinetics up to 1 mmol/Ld-fructose; at higher concentrations the rate ofd-fructose uptake decreased. In the wild-type strain biphasicd-fructose uptake kinetics were observed; the low-affinity component was not found in the mutant, but the high-affinity transport
system persisted. During the exponential phase of growth (ond-glucose) the high-affinityd-fructose system was repressed in the wild-type strain. Mutual competition betweend-fructose andd-glucose as well as the pH dependence of transport of the two hexoses further supported the following conclusion: In the wild-type
strain,d-fructose is taken up both by the specific fructose carrier (K
T=0.22 mmol/L) and the glucose carrier (K
T=9.13 mmol/L). The former does not translocated-glucose, the latter is damaged by the mutation. Finally H+ co-transport and plasma membrane depolarization induced by the onset ofd-fructose transport indicated that the fructose carrier is an H+ symporter. 相似文献
5.
The effects of -aminolaevulinic acid (ALA), porphobilinogen (PBG), -aminobutyric acid (GABA), muscimol, glutamic acid and kainic acid on [3H]2-deoxy-d-glucose uptake by cultured neurons were investigated. Exposure of the cultures for 4 days, to ALA at concentrations as low as 10 M caused a significant, dose-dependent decrease in [3H]2-deoxy-d-glucose uptake. Neither ALA nor PBG appeared to interfere directly with glucose transport into the neuron but 1 mM ALA caused an initial stimulation of [3H]2-deoxy-d-glucose uptake which increased to a maximum after 4 hr and fell to below control values after 19 hr exposure. GABA and muscimol caused similar increases in [3H]2-deoxy-d-glucose uptake but these values remained above control levels after 19 hr exposure. Glutamic acid and kainic acid caused an immediate increase in [3H]2-deoxy-d-glucose uptake which declined to mininum values after 4 hr exposure. The effect of ALA on glucose utilization in neurons may be of particular relevance to patients with acute porphyria where a genetic lesion in neural haem and haemoprotein biosynthesis is postulated to occur. ALA appeared to be more toxic to the neurons than any of the other compounds tested, possibly causing a critical depletion of energy reserves and cell death. 相似文献
6.
Glucose was required for the transport of arabinose into Bifidobacterium breve. The non-metabolisable glucose analogue 2-deoxy-d-glucose (2-DG) did not facilitate assimilation of arabinose. Studies using d-[U-14C]-labelled arabinose showed that it was fermented to pyruvate, formate, lactate and acetate, whereas the principal metabolic products of d-[U-14C]-labelled glucose were acetate and formate. In contrast to glucose, arabinose was not incorporated into cellular macromolecules. A variety of metabolic inhibitors and inhibitors of sugar transport (proton ionophores, metal ionophores, compounds associated with electron transport) were used to investigate the mechanisms of sugar uptake. Only NaF, an inhibitor of substrate level phosphorylation, and 2-DG inhibited glucose assimilation. 2-DG had no effect on arabinose uptake, but NaF was stimulatory. High levels of phosphorylation of glucose and 2-DG by PEP and to a lesser degree, ATP were seen in phosphoenolpyruvate: phosphotransferase (PEP:PTS) assays. These data together with strong inhibition of glucose uptake by NaF suggest a role for phosphorylation in the transport process. Arabinose uptake in B. breve was not directly dependent on phosphorylation or any other energy-linked form of transport but may be assimilated by glucose-dependent facilitated diffusion.Abbreviations (2,4-DNP)
2,4-dinitrophenol
- (2,4-DNP)
carbonylcyanide m-chlorophenylhydrazone
- (CCCP)
(phosphoenolpyruvate phosphotransferase system)
- PEP: PTS
trichloroacetic acid
- (TCA)
2-deoxy-d-glucose
- (2-DG)
2-deoxy-d-glucose 相似文献
7.
Srdjan Novak Tony D'Amore Inge Russell Graham G. Stewart 《Journal of industrial microbiology & biotechnology》1990,6(2):149-155
Summary A number of 2-deoxy-d-glucose (2-DOG) resistant mutants exhibiting resistance to glucose repression were isolated from variousSaccharomyces yeast strains. Most of the mutants isolated were observed to have improved maltose uptake ability in the presence of glucose. Fermentation studies indicated that maltose was taken up at a faster rate and glucose taken up at a slower rate in the mutant strains compared to the parental strains, when these sugars were fermented together. When these sugars were fermented separately, only the 2-DOG resistant mutant obtained fromSaccharomyces cerevisiae strain 1190 exhibited alterations in glucose and maltose uptake compared to the parental strain. Kinetic analysis of sugar transport employing radiolabelled glucose and maltose indicated that both glucose and maltose were transported with higher rates in the mutant strain. These results suggested that the high affinity glucose transport system was regulated by glucose repression in the parental strain but was derepressed in the mutant. 相似文献
8.
d-glucosamine does not serve as a metabolic substrate inSaccharomyces cerevisiae although it stimulates by 15% endogenous respiration. It is taken up by a system or systems shared withd-glucose,d-fructose andd-xylose but apparently not fully with 2-deoxy-d-glucose. Its half-saturation constant is 38±14 mmol/L, in agreement with its inhibitor constant versusd-glucose andd-xylose uptake. Its maximum rate is 69±17 μmol per g dry mass per min. The transport is thermodynamically passive butd-glucosamine distribution follows the membrane potential, reaching ratios of 80∶1 at pH 7.5 and about 1∶1 at pH 4.0. These
rations decrease with increasingd-glucosamine concentration as well as with increasing suspension density, and are affected by metabolic inhibitors. 相似文献
9.
Jürgen Flöge Hilmar Stolte Rolf Kinne 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1984,154(4):355-364
Summary A membrane fraction, rich in brushborder membranes, was prepared from the archinephric duct of the atlantic hagfish (Myxine glutinosa) and the uptake ofd-glucose and other sugars into the membrane vesicles was investigated by a rapid filtration technique. Uptake ofd-glucose was found to be sodium-dependent, phloridzin-inhibitable and osmotically sensitive. A sodium gradient dependent overshoot was demonstrated at 25° C as well as at the more physiological temperature of 4°C. The sodium dependentd-glucose transport was inhibited by -methyl-d-glucoside, but not by 2-deoxy-d-glucose. Furthermore at the same concentration of sugars the initial uptake ofd-glucose was 7.2-fold higher thanl-glucose uptake.d-glucose transport across the membrane in the presence of a sodium gradient was stimulated when SCN– replaced Cl– and inhibited when gluconate replaced Cl–.d-glucose uptake in the presence of a sodium- and potassium gradient was decreased by the addition of valinomycin. In addition, the presence of ad-glucose gradient enhanced sodium uptake into the vesicles as compared to a mannitolgradient. Phloridzin inhibited thed-glucose dependent sodium flux. Thus an electrogenic stereospecific sodium glucose co-transport system, with properties similar to that found in the kidney of higher vertebrates is present in this primitive vertebrate and might participate in secondary-active sugar reabsorption in the archinephric duct. 相似文献
10.
Uptake activities for both glycerol andl-α-glycerol-3-phosphate inPseudomonas aeruginosa strain PAO were induced during growth in the presence of either glycerol ordl-α-glycerol-3-phosphate. Succinate, malate, and glucose exerted catabolite repression control over induction of both uptake
activities. Glycerol uptake exhibited saturation kinetics with an apparentK
m
of 13 μM and aV
max
of 73 nmol/min/mg cell protein. The uptake ofl-α-glycerol-3-phosphate was inhibited by the presence of glycerol, but uptake of glycerol was unaffected by exogenousl-α-glycerol-3-phosphate. Uptake of both substrates by starved, induced cells was stimulated by exogenously providedd-glucose, 2-deoxy-d-glucose,d-gluconate, orl-malate. In a mutant deficient in gluconate uptake and glucose dehydrogenase (EC 1.1.1.47) activities,d-glucose, 2-deoxy-d-glucose, andd-gluconate exerted little or no effect on the uptake of either substrate, butl-malate markedly stimulated the processes. The uptake of both glycerol andl-α-glycerol-3-phosphate, by either starved or unstarved cells, was inhibited by a number of metabolic poisons, including arsenate,
azide, cyanide, 2,4-dinitrophenol, and iodoacetate. 相似文献
11.
K. Yoshioka K. B. Oh M. Saito Y. Nemoto H. Matsuoka 《Applied microbiology and biotechnology》1996,46(4):400-404
A new fluorescent derivative of d-glucose, 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-d-glucose (2-NBDG), which had been previously developed for the analysis of glucose uptake activity by living cells, was investigated to evaluate its applicability for assaying the viability of yeast Candida albicans. Lineweaver-Burk plots showed the uptake of 2-NBDG to be competitively inhibited by d-glucose and not by l-glucose, which suggested the involvement of the glucose transporting system of C. albicans in the uptake of 2-NBDG. A good correlation was obtained between the yeast viability, determined by the plate-count method, and the 2-NBDG uptake activity of yeast cells (correlation constant: r=0.97). This is expected to lead to the development of a new fluorescent probe for the determination of yeast cell viability. 相似文献
12.
Yoshie Terashima 《Mycoscience》1999,40(1):51-56
Mycelial growth of an isolate ofT. bakamatsutake was tested in media with C/N ratio ranging from 0 to 50 and with 32 carbon and 12 nitrogen sources. The isolate grew best
at the C/N ratio of 30. It utilized the monosaccharidesd-glucose,d-mannose, andd-fructose, the disaccharide trehalose, and polysaccharide pectin among the carbon sources; and yeast extract,l-glutamic acid, and ammonium compounds among the nitrogen sources. The growth of ten isolates and secretion of gluconic and
oxalic acids were compared ind-glucose, trehalose, and pectin media. The utilization ofd-glucose, trehalose, and pectin differed among the ten isolates, but all the isolates secreted gluconic acid in thed-glucose media and oxalic acid in the pectin media. 相似文献
13.
John M. Bissonnette M.D. John A. Black Walter K. Wickham Karen M. Acott 《The Journal of membrane biology》1981,58(1):75-80
Summary Glucose uptake into plasma membrane vesicles from the maternal surface of the human placenta was measured with the Millipore filtration technique. Uptake ofd-glucose was dependent on the osmolarity of the incubation medium surrounding the vesicles. Uptake ofd-glucose exceeded that ofl-glucose. The uptake ofd-glucose was not enhanced by placing 100mm NaCl or NaSCN in the medium outside the vesicles (none inside) at the onset of uptake determinations.d-glucose transport was inhibited by cytochalasin B; phloretin, phlorizin, and 1-fluoro-2,4-dinitrobenzene.d-glucose uptake was inhibited by 2-deoxy-d-glucose, 3-O-methyl-d-glucose and to a lesser extent byd-galactose. It was not inhibited by -methyl-d-glucoside. Cytochalasin B binding to the vesicles was 30% inhibited in the presence of 80mm d-glucose. The results indicate that the system for facilitated transport ofd-glucose at the maternal face of the placenta is distinctly different from that on the brush-border membrane of intestine or renal tubule and more closely resembles that of human erythrocyte. 相似文献
14.
Stationary unaerated cells ofS. pombe containing endogenous substrates but not energized by any exogenous ones take up 2-deoxy-d-glucose, 6-deoxy-d-glucose,d-xylose andd-arabinose actively over diffusion equilibrium. The active uptake is inhibited by 20–100 mmol/L H2O2 which causes an increase inK
T but has no effect onJ
max. This “competitive inhibition” indicates that H2O2 affects directly the sugar binding sites of the transporters. The ATP-binding site of the plasma membrane H+-ATPase is also affected by 100 mmol/L H2O2; theK
T decreases 7-fold,J
max about 2.5-fold. These effects are not likely to be mediated by membrane lipid peroxidation which appears to be lacking inS. pombe, and this lack may be one of the reasons for the high resistance of this yeast to H2O2. Because of thisS. pombe represents a suitable system for studying direct effects of oxidants on membrane proteins. 相似文献
15.
A novel phosphorylase from Clostridium phytofermentans belonging to the glycoside hydrolase family (GH) 65 (Cphy1874) was characterized. The recombinant Cphy1874 protein produced
in Escherichia coli showed phosphorolytic activity on nigerose in the presence of inorganic phosphate, resulting in the release of d-glucose and β-d-glucose 1-phosphate (β-G1P) with the inversion of the anomeric configuration. Kinetic parameters of the phosphorolytic activity
on nigerose were k
cat = 67 s−1 and K
m = 1.7 mM. This enzyme did not phosphorolyze substrates for the typical GH65 enzymes such as trehalose, maltose, and trehalose
6-phosphate except for a weak phosphorolytic activity on kojibiose. It showed the highest reverse phosphorolytic activity
in the reverse reaction using d-glucose as the acceptor and β-G1P as the donor, and the product was mostly nigerose at the early stage of the reaction. The
enzyme also showed reverse phosphorolytic activity, in a decreasing order, on d-xylose, 1,5-anhydro-d-glucitol, d-galactose, and methyl-α-d-glucoside. All major products were α-1,3-glucosyl disaccharides, although the reaction with d-xylose and methyl-α-d-glucoside produced significant amounts of α-1,2-glucosides as by-products. We propose 3-α-d-glucosyl-d-glucose:phosphate β-d-glucosyltransferase as the systematic name and nigerose phosphorylase as the short name for this Cphy1874 protein. 相似文献
16.
Summary A membrane extract enriched with the Na+-dependentd-glucose transport system was obtained by differential cholate solubilization of rat renal brush border membranes in the presence of 120mm Na+ ions. Sodium ions were essential in stabilizing the transport system during cholate treatment. This membrane extract was further purified with respect to its Na+-coupledd-glucose transport activity and protein content by the use of asolectin-equilibrated hydroxylapatite. The reconstituted proteoliposomes prepared from this purified fraction showed a transient accumulation ofd-glucose in response to a Na+ gradient. The observed rate of Na+-coupledd-glucose uptake by the proteoliposomes represented about a sevenfold increase as compared to that of the reconstituted system derived from an initial 1.2% cholate extract of the membranes. Other Na+-coupled transport systems such asl-alanine, -ketoglutarate and phosphate were not detected in these reconstituted proteoliposomes. 相似文献
17.
Brush border membrane vesicles (BBMV) enriched in sucrase, maltase and alkaline phosphatase, and impoverished in Na+-K+-ATPase, were isolated from proximal and distal intestine of the gilthead sea bream (Sparus aurata) by a MgCl2 precipitation method. Vesicles were suitable for the study of the characteristics of D-glucose apical transport. Only one D-glucose carrier was found in vesicles from each intestinal segment. In both cases, the D-glucose transport system was sodium-dependent, phlorizin-sensitive, significantly inhibited by D-glucose, D-galactose, α-methyl-D-glucose, 3-O-methyl-D-glucose and 2-deoxy-D-glucose, and showed stereospecificity. Apparent affinity constants of D-glucose transport (Kt) were 0.24 ± 0.03 mM in proximal and 0.18 ± 0.03 mM in distal intestine. Maximal rate of influx (Jmax) was 47.3 ± 2.2 pmols. mg−1 protein for proximal and 27.3 ± 3.6 pmols. mg−1 protein for distal intestine. Specific phlorizin binding and relative abundance of an anti-SGLT1 reactive protein were significantly higher in proximal than in distal BBMV. These results suggest the presence of the same D-glucose transporter along the intestine, with a higher density in the proximal portion. This transporter is compatible with the sodium-dependent D-glucose carrier described for other fish and with the SGLT1 of higher vertebrates.This revised version was published online in June 2005 with a corrected cover date. 相似文献
18.
Schizosaccharomyces pombe cells grow on d-gluconate as the sole carbon and energy source. d-Gluconate is taken up in symport with protons by a specific symporter, pH being the sole driving force. d-Gluconate uptake is independent of the sugar transporting system (e.g. for d-glucose) and of . The carrier is expressed constitutively, and its activity is not subject to glucose repression. Hence, d-gluconate is a suitable carbon and energy source for growth, when d-glucose or other hexoses have to be eliminated e.g. for selection of mutants deficient in hexose transport.Abbreviations 2-DG
2-deoxy-d-glucose
- CCCP
carbonylcyanide m-chlorophenylhydrazone
- pH
pH-gradient
-
electrical potential difference across the plasma membrane
- SD
standard deviation
- SEM
standard error of the mean
- TPP+
tetraphenylphosphonium 相似文献
19.
Summary Glucose transport was studied in marine mussels of the genusMytilus. Initial observations, with intact animals and isolated gills, indicated that net uptake of glucose occurred in mussels by a carrier-mediated, Na+-sensitive process. Subsequent studies included use of brush-border membrane vesicles (BBMV) in order to characterize this transport in greater detail. The highest activity of Na+-dependent glucose transport was found in the brush-border membrane fractions used in this study, while basal-lateral membrane fractions contained the highest specific binding of ouabain. Glucose uptake into BBMV showed specificity for Na+, and concentrative glucose transport was observed in the presence of an inwardly directed Na+ gradient. There was a single saturable pathway for glucose uptake, with an apparentK
t of 3 m in BBMV and 9 m in intact gills. The kinetics of Na+ activation of glucose uptake were sigmoidal, with apparent Hill coefficients of 1.5 in BBMV and 1.2 in isolated gills, indicating that more than one Na+ may be involved in the transport of each glucose. Harmaline inhibited glucose transport in mussel BBMV with aK
i of 44 m. The uptake of glucose was electrogenic and stimulated by an inside-negative membrane potential. The substrate specificity in intact gills and BBMV resembled that of Na+-glucose cotransporters in other systems;d-glucose and -methyl glucopyranoside were the most effective inhibitors of Na+-glucose transport,d-galactose was intermediate in its inhibition, and there was little or no effect ofl-glucose,d-fructose, 2-deoxy-glucose, or 3-O-methyl glucose. Phlorizin was an effective inhibitor of Na+-glucose uptake, with an apparentK
i of 154nm in BBMV and 21nm in intact gills. While the qualitative characteristics of glucose transport in the mussel gill were similar to those in other epithelia, the quantitative characteristics of this process reflect adaptation to the seawater environment of this animal. 相似文献
20.
Muller AP Rotta LN Kawano C Leszczinski DN Schweigert ID Londero LG Gravina FS da Silveira CK de Souza CG Battu CE Gonçalves CA de Souza DO Perry ML 《Neurochemical research》2006,31(3):417-422
We studied the effect of different concentrations of 2-deoxy-d-glucose on the l-[U-14C]leucine, l-[1-14C]leucine and [1-14C]glycine metabolism in slices of cerebral cortex of 10-day-old rats. 2-deoxy-d-glucose since 0.5 mM concentration has inhibited significantly the protein synthesis from l-[U-14C]leucine and from [1-14C]glycine in relation to the medium containing only Krebs Ringer bicarbonate. Potassium 8.0 mM in incubation medium did not stimulate the protein synthesis compared to the medium containing 2.7 mM, and at 50 mM diminishes more than 2.5 times the protein synthesis compared to the other concentration. Only at the concentration of 5.0 mM, 2-deoxy-d-glucose inhibited the CO2 production and lipid synthesis from l-[U-14C] leucine. This compound did not inhibit either CO2 production, or lipid synthesis from [1-14C]glycine. Lactate at 10 mM and glucose 5.0 mM did not revert the inhibitory effect of 2-deoxy-d-glucose on the protein synthesis from l-[U-14C]leucine. 2-deoxy-d-glucose at 2.0 mM did not show any effect either on CO2 production, or on lipid synthesis from l-[U-14C]lactate 10 mM and glucose 5.0 mM. 相似文献