共查询到20条相似文献,搜索用时 31 毫秒
1.
Alexander P.R. Theuvenet Willem M.H. van de Wijngaard George W.F.H. Borst-Pauwels 《生物化学与生物物理学报:生物膜》1983,730(2):255-262
The uptake of 9-aminoacridine is studied in the yeast Saccharomyces cerevisiae by fluorescence and absorbance measurements of the dye. Uptake of the dye proceeds via two pathways. One pathway consists of a diffusion of the non-protonated form. At high pH (7.5) this pathway is the predominant one, and the dye distributes between the cell inner and the medium according to the ratio of the proton concentrations in the two compartments. In other words, at high pH 9-aminoacridine behaves as a probe of the H+ gradient across the yeast cell membrane. At low external pH (4.5) a second pathway is involved. Much greater accumulation ratios for the dye are observed than can be accounted for by the H+ gradient across the membrane. The transport system predominantly responsible for the great accumulation of the dye appears to be inducible, to require metabolic energy and to be saturable. This transport system is competitively inhibited by thiamine, and also by dibenzyldimethylammonium and thiaminedisulfide, two specific inhibitors of the thiamine carrier in the yeast. On the other hand, the thiamine uptake by the yeast cells is competitively inhibited by 9-aminoacridine. In addition, uptake of 9-aminoacridine is greatly reduced in the thiamine transport-negative mutant of S. cerevisiae, PT-R2. It is concluded that at low pH 9-aminoacridine is taken up by yeast via the thiamine carrier of the cell and that, consequently, the dye may be applied as a probe of this transport system. 相似文献
2.
Hartmannella culbertsoni trophozoites are transformed into viable cysts on exposure to a non-nutrient agar medium containing 15 mM MgCl2 and 20 mM taurine. Amebae differentiating in this encystment medium incorporate more uracil-2-14C into RNA and more leucine-1-14C or valine-1-14C into proteins than controls. Encysting organisms incorporate significantly more glucose-U-14C into cellulose and glucosamine-1-14C into mucopolysaccharides. Incorporation of glucose-U-14C into cellulose and of glucosamine-1-14C into mucopolysaccharides are inhibited by actinomycin D or cycloheximide. 相似文献
3.
Summary Mutual correction of co-cultivated fibroblasts from patients with Hunter's and Hurler's syndrome could be inhibited by either fructose 1-phosphate or mannose 6-phosphate. In the presence of fructose 1-phosphate a 50% mixture of fibroblasts from a patient with Hunter's syndrome and a normal homozygous individual showed an increased35S-sulphate incorporation into acid mucopolysaccharides. When fibroblast cultures from one obligate and two possible carriers of Hunter's syndrome were tested for35S-sulphate incorporation, the cultures showed either twice the normal35S-sulphate incorporation into acid mucopolysaccharides in the presence of fructose 1-phosphate or an abnormally high incorporation in the presence as well as in the absence of the sugar phosphate. 相似文献
4.
Acid mucopolysaccharide synthesis in the secondary palate of the developing rat at the time of rotation and fusion 总被引:5,自引:0,他引:5
The amount of hexosamines and acid mucopolysaccharides present in the rat secondary palate increases during the critical stages of palatogenesis, namely, rotation and fusion. The synthesis of acid mucopolysaccharides in vivo and in vitro in the palate was determined by the incorporation of 3H-glucosamine and Na2S35O4. The labeled mucopolysaccharides were isolated by DEAE-cellulose chromatography and were identified on the basis of several criteria as hyaluronic acid and sulfated acid mucopolysaccharides. Hyaluronic acid accounted for approximately 60% of the total acid mucopolysaccharides synthesized in the palate both in vivo and in vitro. DON (6-diazo-5-oxonorleucine), a known inhibitor of acid mucopolysaccharide synthesis, inhibited the incorporation of 3H-glucosamine and Na2S35O4 by palatal shelves in vitro by 70%. 相似文献
5.
Ruthenium red, a powerful inhibitor of Ca2+ transport by mitochondria, does not inhibit the active Ca2+ uptake by sarcoplasmic reticulum isolated from rabbit skeletal muscle promoted by 5 mM ATP-Mg in the presence or absence of potassium oxalate. Although concentrations of ruthenium red up to 100 μM do not affect the active uptake of Ca2+, 25 μM of the inorganic dye inhibit the passive binding of Ca2+ by about 50%. This inhibitory effect is observed in sarcoplasmic reticulum even after its lipid fraction is extracted with acetone.Although active Ca2+ uptake by sarcoplasmic reticulum is not inhibited by ruthenium red, in the absence of oxalate it inhibits significantly the Ca2+-dependent ATPase activity but not the Mg2+-ATPase. However, if potassium oxalate is present, the Ca2+-stimulated ATPase is not sensitive to the dye. It is not clear how oxalate functions to protect the Ca2+-ATPase against the inhibitor effect of ruthenium red.The high sensitivity to ruthenium red of the Ca2+ transport mechanism in mitochondria as compared to the Ca2+ transport in sarcoplasmic reticulum may be useful in determining the extent to which each organelle functions in the cell to regulate intracellular free Ca2+. 相似文献
6.
The sensitivity of the fluorescent dye, 3,3′-diethylthiadicarbocyanine (DiS-C2(5)), was too low for the detection of membrane potential changes in rat small intestinal membrane vesicles. Only after adding LaCl3 or after fractionation of the intestinal membranes by free-flow electrophoresis could the dye be used to monitor electrogenic Na+-dependent transport systems. It is concluded that the response of this potential-sensitive dye is influenced by the negative surface charge density of the vesicles. 相似文献
7.
The hydrophyllic dyes fluorescein glutamic acid, fluorescein glutamylglutamic acid (F(Glu)2), fluorescein hexaglycine, fluorescein leucyldiglutamyl-leucine and 6-carboxyfluorescein are unable to pass the plasmalemma in leaves of E. densa. However, when injected into single cells the dye conjugates of molecular weight 665 dalton or less move freely from cell-to-cell. This intercellular movement presumably occurs via the plant symplast. Movement of F(Glu)2 from the injected cell occurs with greatly reduced frequency when Ca2+, Mg2+ or Sr2+ are injected into the cell immediately prior to the dye. The fraction of dye injections leading to movement declines with increasing group II ion concentration in the electrode tip, up to 10 mM. Sodium and K ions do not affect dye movement. When dye injection is delayed 30 min after Ca2+ injection, dye movement is no longer inhibited. Thus the cells recover from the Ca2+ injection, indicating that the ion does not cause major cell damage. Recovery from Mg2+ injection is not complete within 60 min. Treatment of leaves with chemicals expected to raise the concentration of free intracellular group II ions, notably the mitochondrial uncoupler carbonyl cyanide p-trifluoromethoxyphenyl hydrazone, the inhibitor of mitochondrial Ca2+ uptake trifluralin, or the ionophore A23187 also inhibits dye movement, while the calmodulin inhibitor trifluoperazine does not. Cytoplasmic streaming is inhibited by Ca2+ or Mg2+ injection and by the metabolic inhibitors. However when streaming is stopped by cytochalasin B, dye movement is not inhibited. Hence steaming is not necessary for dye movement. Thus the cytoplasmic concentration of free group II ions may directly regulate the permeability of the plant symplast. 相似文献
8.
Tetsuro Mohri Hitomi Sasaki 《Biochemical and biophysical research communications》1982,108(2):854-860
Pretreatment(2 h) of Chang liver cells with glucagon and dibutyryl-cAMP has been shown to stimulate specifically the Na+-independent transport system for amino acids. The stimulation of Na+-independent leucine uptake was completely or largely inhibited by prior incubation(30 min) of the cells with colchicine or cytochalasin B. Glucagon treatment colud only change the Vmax value of the high-affinity component of Na+-independent leucine transport and the apparent Km value of the low-affinity one, whereas dibutyryl-cAMP treatment more or less affected all of the kinetic constants of both of the two components in favor of promotion of leucine tansport. These results of kinetic analysis indicate that dibutyryl-cAMP may not perfectly play the role of glucagon in activation of the Na+-independent transport system for leucine. 相似文献
9.
Summary Thel-alanine-dependent transport of sodium ions across the plasma membrane of rat-liver parenchymal cells was studied using isolated plasma membrane vesicles. Sodium uptake is stimulated specifically by thel-isomer of alanine and other amino acids, whose transport is sodium-dependent in rat-liver plasma membrane vesicles. Thel-alanine-dependent sodium flux across the membrane is inhibited by an excess of Li+ ions, but not by K+ or choline ions. Sodium transport is sensitive to-SH reagents and ionophores, and is an electrogenic process: a membrane potential (negative inside) can enhancel-alanine-dependent sodium accumulation. The data presented provide further evidence for a sodium-alanine cotransport mechanism. 相似文献
10.
The system for uptake of maltose in Pseudomonas fluorescens W was inducible. Using a mutant strain unable to hydrolyze maltose, it was shown that maltose was taken up unaltered against a concentration gradient. Uptake of 14C maltose was only significantly inhibited by nonradioactive maltose or maltotriose. These were the only sugars that could displace accumulated radioactive maltose in the strain unable to hydrolyze maltose. Uptake exhibited saturation kinetics and was inhibited by energy poisons, indicating that this system was one of active transport. Sulfhydryl-binding reagents reversibly inhibited maltose uptake. No transport ability was lost when cells were subjected to osmotic shock. Using the protein-binding dye 7-diazonium-1,3-naphthalene disulfonate a protein or proteins located in or external to the cell membrane was implicated in maltose transport. The hydrolysis of p-nitrophenyl--D-glucoside (PNPG) was used as an indirect measure of transport ability since penetration of PNPG, not its hydrolysis, was the rate-limiting step.Abbreviations PNPG
paranitrophenyl--D-glucoside
- NDS
7-diazonium-1,3-naphthalene disulfonic acid
- PMB
p-hydroxymercuribenzoate
- MBS
p-chloromercuriphenylsulfonic acid
- PCMB
p-chloromercuribenzoate
- CCCP
carbonyl cyanide m-chlorophenylhydrazone
- DNP
2,4-dinitrophenol
- HOQNO
2-heptyl-4-hydroxyquinoline-N-oxide 相似文献
11.
《生物化学与生物物理学报:生物膜》1992,1110(1):123-126
Monitoring the fluorescence quenching of the pH-sensitive dye Acridine orange, proton accumulation in the presence of an inside-negative transmembrane potential was measured in eel (Anguilla anguilla) intestinal brush-border membrane vesicles. It was demonstrated that the proton accumulation was specifically increased by the presence of the dipeptide glycyl-glycine in the extravesicular space, showing saturation kinetics at increasing dipeptide concentrations and was specifically inhibited by diethylpyrocarbonate. Data reported suggest the presence of an electrical-potential-dependent H+/glycyl-glycine cotransport system in the eel intestinal brush-border membrane vesicles. 相似文献
12.
The fluorescent dye 10-N-nonyl acridine orange (NAO), known as specifically associated with mitochondria, has been reported to have a cytotoxic effect when high doses were applied to cells. Presently, the biochemical basis of its toxicity was investigated on isolated rat liver mitochondria. At low concentrations, NAO strongly inhibited state 3 respiration and ATP synthesis. At high concentrations, electron transport, ATP hydrolysis, Pi-transport and adenine nucleotide activities were also decreased. All these inhibitions can be explained by probe-cardiolipin interactions which could induce the collapse of energy conversion and/or the modification of membrane fluidity. 相似文献
13.
The interaction of sulfated mucopolysaccharides and lectins has been studied by determining the amount of precipitate formed when mucopolysaccharides are added to a solution of concanavalin A or a partially purified lectin preparation from red kidney bean (Phaseolus vulgaris). The amount of insoluble complex obtained when a given mucopolysaccharide is added to a solution of partially purified red kidney bean preparation is pH dependent. The reaction of concanavalin A and heparin has also been studied by adding increasing amounts of mucopolysaccharide to a fixed amount of lectin. This interaction results in the development of a precipitin-like curve and leads to the isolation of a heparin fraction which has been found to be more reactive with respect to formation of a precipitate than the original heparin preparation. Monosaccharides such as α-methyl-d-mannopyranoside and N-acetyl-d-glucosamine which are known to bind specifically to the lectin, greatly inhibit precipitate formation. The interactions between sulfated mucopolysaccharides and lectins have been used to isolate various sulfated mucopolysaccharides. 相似文献
14.
The effect of the transport of tricarboxylic acid cycle intermediates on the membrane potential of renal brush border vesicles was studied using fluorescence of the cyanine dye, 3,3′-dipropylthiadicarbocyanine iodide. The behavior of the dye in the preparation was established with valinomycin-induced K+-diffusion potentials; increases in fluorescence were associated with depolarizing conditions. Addition of 1 mM succinate or citrate to membrane/dye suspensions produced transient increases in fluorescence, indicative of a depolarizing event(s) associated with the transport of these substrates. The transient response in fluorescence was Na+ dependent, of greater magnitude under Na+-gradient as compared to Na+-equilibrium conditions, and was a saturable function of substrate concentration. The specificity of the fluorescence response was identical to that obtained from studies of the competitive inhibition of succinate transport by tricarboxylic acid cycle intermediates and analogs. We conclude that the major tricarboxylic acid cycle intermediates are transported via a common Na+-dependent transport system in renal brush border membranes. 相似文献
15.
Inhibition of casein kinase II by heparin 总被引:24,自引:0,他引:24
Casein kinase II, a cyclic nucleotide-independent protein kinase from rabbit reticulocytes, was shown to be inhibited by heparin. Heparin specifically inhibited the enzyme and had no effect on other protein kinases, including casein kinase I, the type I and II cAMP-dependent protein kinases, protease-activated kinase I, and the hemin-controlled repressor. Heparan sulfate was found to be 40-fold less effective than heparin towards casein kinase II; other acid mucopolysaccharides had little or no effect on the enzymatic activity. Steady state studies revealed that heparin acted as a competitive inhibitor with respect to the substrate, casein. A value of 20 ng/ml or about 1.4 nM was obtained for the apparent Ki. The inhibition was not reversed by ATP and varying the ATP and heparin concentrations in the assay only altered the maximum velocity. 相似文献
16.
M J Murphy D A Dunbar F P Guengerich L S Kaminsky 《Archives of biochemistry and biophysics》1981,212(2):360-369
The effects of ethanol and acetaldehyde on rat intestinal microvillus membrane integrity and glucose transport function were examined in vitro with purified membrane vesicles. Ethanol could influence glucose transport function by alterations in the conformation of the carrier, the lipid environment surrounding the carrier, or in the transport driving force (Na+ electrochemical gradient). Due to the rapid nature of glucose uptake, transport was assayed with the use of an apparatus that permitted uptake measurements as early as 1 s. Ethanol (340 mm) partially and acetaldehyde (44 mm) completely inhibited concentrative glucose uptake throughout the 1-min time course. Their inhibitory effects were reversible and irreversible, respectively. Kinetic measurements made during the initial rate of uptake (at 2 s) with various concentrations of glucose (0.05–8 mm) showed that ethanol and acetaldehyde both caused a decrease in V. Although ethanol did not substantially alter the transport Km, acetaldehyde increased the Km almost 50%. To determine whether ethanol or acetaldehyde directly interfered with glucose carrier function, uptake was measured in the presence of equilibrated Na+. Only acetaldehyde had a significant inhibitory effect under these conditions. Membrane permeability, as determined by efflux of preloaded 6-carboxyfluorescein dye, increased upon exposure of the vesicles to ethanol or acetaldehyde. Membrane fluidity measurements by fluorescence polarization showed that only acetaldehyde had a significant fluidizing effect. These results indicate that ethanol and acetaldehyde acted to perturb membrane integrity and inhibited glucose uptake indirectly by allowing the Na+ gradient to dissipate. Acetaldehyde, which had a stronger inhibitory effect than ethanol, appeared also to directly inhibit carrier function. 相似文献
17.
Helminthosporium maydis T Toxin Decreased Calcium Transport into Mitochondria of Susceptible Corn 总被引:7,自引:6,他引:1
下载免费PDF全文
![点击此处可从《Plant physiology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
The effects of purified Helminthosporium maydis T (HmT) toxin on active Ca2+ transport into isolated mitochondria and microsomal vesicles were compared for a susceptible (T) and a resistant (N) strain of corn (Zea mays). ATP, malate, NADH, or succinate could drive 45Ca2+ transport into mitochondria of corn roots. Ca2+ uptake was dependent on the proton electrochemical gradient generated by the redox substrates or the reversible ATP synthetase, as oligomycin inhibited ATP-driven Ca2+ uptake while KCN inhibited transport driven by the redox substrates. Purified native HmT toxin completely inhibited Ca2+ transport into T mitochondria at 5 to 10 nanograms per milliliter while transport into N mitochondria was decreased slightly by 100 nanograms per milliliter toxin. Malate-driven Ca2+ transport in T mitochondria was frequently more inhibited by 5 nanograms per milliliter toxin than succinate or ATP-driven Ca2+ uptake. However, ATP-dependent Ca2+ uptake into microsomal vesicles from either N or T corn was not inhibited by 100 nanograms per milliliter toxin. Similarly, toxin had no effect on proton gradient formation ([14C]methylamine accumulation) in microsomal vesicles. These results show that mitochondrial and not microsomal membrane is a primary site of HmT toxin action. HmT toxin may inhibit formation of or dissipate the electrochemical proton gradient generated by substrate-driven electron transport or the mitochondrial ATPase, after interacting with a component(s) of the mitochondrial membrane in susceptible corn. 相似文献
18.
Different methods to estimate the plasma membrane potential difference (PMP) of yeast cells with fluorescent monitors were
compared. The validity of the methods was tested by the fluorescence difference with or without glucose, and its decrease
by the addition of 10 mM KCl. Low CaCl2 concentrations avoid binding of the dye to the cell surface, and low CCCP concentrations avoid its accumulation by mitochondria.
Lower concentrations of Ba2+ produce a similar effect as Ca2+, without producing the fluorescence changes derived from its transport. Fluorescence changes without considering binding
of the dyes to the cells and accumulation by mitochondria are overshadowed by their distribution between this organelle and
the cytoplasm. Other factors, such as yeast starvation, dye used, parameters of the fluorescence changes, as well as buffers
and incubation times were analyzed. An additional approach to measure the actual or relative values of PMP, determining the
accumulation of the dye, is presented. 相似文献
19.
Anne Kearns Louise Cole Chris R. Hawes David E. Evans 《Plant Physiology and Biochemistry》1998,36(12):879-887
Uptake of the fluorescent dye hydroxypyrenetrisulfonate by carrot (Daucus carota L.) suspension cells only occurs when the external (medium) pH falls to below 4.0. Uptake of the dye was shown to be inhibited by a range of compounds, including the ‘Good’ buffers MES, HEPES, HEPPSO and HEPPS, the growth medium component coconut water (CW) and probenecid, an organic anion translocator inhibitor. Tris(hydroxymethyl)aminomethane stimulated transport. Buffer effects were not correlated with pKa values. Inhibitors of dye uptake (MES, probenecid, CW) also inhibited medium acidification by the cells and Tris stimulated acidification and uptake. Uptake of HPTS correlated strongly with external pH and was restored when external pH was experimentally reduced to below 4.0 even in the presence of inhibitors. This suggests that inhibitors of HPTS uptake at the plasma membrane act primarily by preventing the establishment of a low external pH required for transport. The implications of this on the mechanism of HPTS transport are discussed. 相似文献
20.
Addition of different concentrations of heavy metal ions (Hg2+, Cu2+, Ni2+ and Pb2+) inhibited the photosystem 2 catalyzed electron transport activity (H2O→p-benzo-quinone) of the cyanobacteriumSpirulina platensis. Hg2+ caused the inhibition in electron transport activity in very low concentrations compared to the other metal ions. Hg2+ at this low concentration specifically altered the spectral properties of phycocyanin of the phycobilisomes in the intact cells ofSpirulina, whereas other heavy metal ions were ineffective in this sense. 相似文献