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Anastacio M. Hoyumpa Sharon Nichols Steven Schenker Frederick A. Wilson 《生物化学与生物物理学报:生物膜》1976,436(2):438-447
As part or a systematic study of alcoholism and thiamine absorption, the effect of diet-induced thiamine deficiency and the role of the unstirred water layer on thiamine transport were investigated. Using 3H-labeled dextran as a marker of adherent mucosal volume, jejunal uptake of 14C-labeled thiamine hydrochloride was measured, in vitro, in thiamine-deficient rats and pair-fed controls. Uptake of low thiamine concentrations (0.2 and 0.5 μM) was greater in the thiamine-deficient rats thatn in the controls. In contrast, uptake rates for high thiamine concentrations (20 and 50 μM) were similar in both groups. While was unaltered, was decreased in thiamine deficiency, suggesting a decrease in unstirred water layer thickness. Accordingly, the thickness of the water layer was measured in both groups of animals and correlated with and under unstirred and st irred conditions. Without stirring, there was no difference in between the two groups. In contrast, both and the water layer were reduced in the thiamine-deficient rats. With stirring, was not affected, but both and the water layer thickness were reduced to similar values in both groups. Reversal of thiamine deficiency resulted in the return of thiamine uptake and the unstirred water layer thickness to control values. These data support the concept of a dual system of thiamine transport and emphasize the role of the unstirred water layer as an important determinant of transport kinetics not only under physiologic situations but also in diet-induced rat thiamine deficiency, a model for a clinical pathological state. The decrease in the unstirred water layer thickness in thiamine deficiency may be also viewed as a possible adaptive mechanism to facilitate absorption of meager supplies of thiamine. 相似文献
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Sulfate uptake by Saccharomyces cerevisiae is stimulated about 12-fold by preincubation of cells with 1% d-glucose or 1% ethanol. The remains unchanged (0.34–0.38 mM), the increase from 18–20 to 195–230 and 170–185 nmol/min per g dry wt., respectively, after glucose and ethanol preincubation. The stimulation involves protein synthesis (it is suppressed by cycloheximide), has a half-time of 18 min and requires mitochondrial respiration (no or low effect in respiration-deficient mutants and those lacking ADP-ATP transport in mitochondria, as well as after anaerobic preincubation of the wild-type strain, and in low-phosphate cells). The presence of NH4+ and some amino acids (e.g., leucine, aspartate, cysteine and methionine) depressed the stimulation while that of cationic amino acids (typically arginine and lysine) and of K+ increased it by 50–80%. The stimulated (i.e., newly synthesized) transport system was degraded with a half-life of about 10 min. 相似文献
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The transport of sucrose by selected mutant and wild-type cells of Streptococcus mutans was studied using washed cocci harvested at appropriate phases of growth, incubated in the presence of fluoride and appropriately labelled substrates. The rapid sucrose uptake observed cannot be ascribed to possible extracellular formation of hexoses from sucrose and their subsequent transport, formation of intracellular glycogen-like polysaccharide, or binding of sucrose or extracellular glucans to the cocci. Rather, there are at least three discrete transport systems for sucrose, two of which are phosphotransferases with relatively low apparent values and the other a non-phosphotransferase (non-PTS) third transport system (termed TTS) with a relatively high apparent . For strain 6715-13 mutant 33, the values are 6.25·10?5 M, 2.4·10?4 M, and 3.0·10?3 M, respectively; for strain NCTC-10449, the values are 7.1·10?5 M, 2.5·10?4 M and 3.3·10?3 M, respectively. The two lower systems could not be demonstrated in mid-log phase glucose-adapted cocci, a condition known to repress sucrose-specific phosphotransferase activity, but under these conditions the highest system persists. Also, a mutant devoid of sucrose-specific phosphotransferase activity fails to evidence the two high affinity (low apparent ) systems, but still has the lowest affinity (highest ) system. There was essentially no uptake at 4°C indicating these processes are energy dependent. The third transport system, whose nature is unknown, appears to function under conditions of sucrose abundance and rapid growth which are known to repress sucrose-specific phosphotransferase activity in S. mutans. These multiple transport systems seem well-adapted to S. mutans which is faced with fluctuating supplies of sucrose in its natural habitat on the surfaces of teeth. 相似文献
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We determine the kinetic parameters and of lactose transport in Escherichia coli cells as a function of the electrical potential difference (Δψ) at pH 7.3 and . We report that transport occurs simultaneously via two components: a component which exhibits a high (larger than 10 mM) and whose contribution is independent of Δψ, a component which exhibits a low independent of Δψ (0.5 mM) but whose increases drastically with increasing Δψ. We associate these components of lactose transport with facilitated diffusion and active transport, respectively. We analyze the dependence upon Δψ of and of the active transport component in terms of a mathematical kinetic model developed by Geck and Heinz (Geck, P. and Heinz, E. (1976) Biochim. Biophys. Acta 443, 49–63). We show that within the framework of this model, the analysis of our data indicates that active transport of lactose takes place with a H+/lactose stoichiometry greater than 1, and that the lac carrier in the absence of bound solutes (lactose and proton(s)) is electrically neutral. On the other hand, our data relative to facilitated diffusion tend to indicate that lactose transport via this mechanism is accompanied by a H+/lactose stoichiometry smaller than that of active transport. We discuss various implications which result from the existence of H+/lactose stoichiometry different for active transport and facilitated diffusion. 相似文献
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Alanine uptake by rabbit ileal mucosa has been measured in the presence and absence of Na+ to establish the characteristics of biological variation. Common values, calculated for two systems of alanine entry, were used for estimation of two Jmax values for individual rabbits. The distribution of and within the population was normal and there was minimal interaction between these two parameters, judged by a cross-correlation analysis. Ways of processing data to account for this type of animal variation are discussed. 相似文献
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Adenosine triphosphate-dependent glutathione transport was characterized using inside-out vesicles made from human erythrocytes. Kinetic analysis of the glutathione disulfide (GSSG) transport showed a biphasic Line-weaver-Burk plot as a function of GSSG concentration suggesting the operation of two different processes. One phase had a high affinity for GSSG and a low transport velocity. Most active at acidic pH and at 25°C, this transport activity was easily lost during the storage of vesicles at 4°C. The for Mg-ATP was 0.63 mM; guanosine triphosphate (GTP) substituted for ATP gave a 340% stimulation of transport activity. Neither dithiothreitol nor thiol reagents affected this transport process. The other phase had a low affinity for GSSG and a high transport velocity. Most active at pH 7.2 and 37°C, this transport activity was stable during storage of vesicles at 4°C for several days. The for Mg-ATP was 1.25 mM; GTP substituted with no change in activity. Dithiothreitol increased the V but did not alter the , and thiol reagents inhibited the transport. These findings suggest that there are two independent transfer processes for GSSG in human erythrocytes. 相似文献
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The Na+-independent leucine transport system is resolved into two components by their different affinity ( about 44 μM and 8.0 mM) for leucine in the Chang liver cell. Treatment of the cells with (1 mM) specifically stimulates the high-affinity component of the Na+-independent system by greatly increasing its value, whereas the value of the low-affinity component is markedly lowered. The stimulatory effect of on leucine transport is reduced by prior treatment of the cells with 2,4-dinitrophenol, but this phenomenon seems to be irrelevant to the ATP-depleting action of the uncoupler. The treatment with 2,4-dinitrophenol has been found not to be inhibitory on the subsequent Na+-independent leucine uptake itself. Treatment with dibucaine, a phospholipid-interacting drug, also reduces to varying degrees (depending on its concentration) the stimulatory effect of on the subsequent leucine uptake, although pretreatment with dibucaine can stimulate the Na+-independent leucine uptake itself. We conclude that the stimulatory effect of on leucine transport is not correlated with the energy level of cell, but involves the perturbation of the membrane bilayer structures. 相似文献
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The effect of pH on the kinetic parameters for the chloroperoxidase-catalyzed N-demethylation of N,N-dimethylaniline supported by ethyl hydroperoxide was investigated from pH 3.0 to 7.0. Chloroperoxidase was found to be stable throughout the pH range studied. Initial rate conditions were determined throughout the pH range. The Vmax for the demethylation reaction exhibited a pH optimum at approximately 4.5. The Km for N,N-dimethylaniline increased with decreasing pH, while the Km for ethyl hydroperoxide varied in a manner paralleling Vmax. Comparison of the values for N,N-dimethylaniline and ethyl hydroperoxide indicated that the interaction of N,N-dimethylaniline with chloroperoxidase compound I was rate-limiting below pH 4.5, while compound I formation was rate-limiting above pH 4.5. The log of the for ethyl hydroperoxide was independent of pH, indicating that chloroperoxidase compound I formation is not affected by ionizations in this pH range. The plot of the log of the for N,N-dimethylaniline versus pH indicated an ionization on compound I with a pK of approximately 6.8. The plot of the log of the Vmax versus pH indicated an ionization on the compound I-N,N-dimethylaniline complex, with a pK of approximately 3.1. The results show that chloroperoxidase can demethylate both the protonated and neutral forms of N,N-dimethylaniline (pK approximately 5.0), suggesting that hydrophobic binding of the arylamine substrate is more important in catalysis than ionic bonding of the amine moiety. For optimal catalysis, a residue in the chloroperoxidase compound I-N,N-dimethylaniline complex with a pK of approximately 3.1 must be deprotonated, while a residue in compound I with a pK of approximately 6.8 must be protonated. 相似文献
10.
Diethelm Kleiner 《生物化学与生物物理学报:生物膜》1982,688(3):702-708
Klebsiella pneumoniae can accumulate methylammonium up to 80-fold by means of a transport system as indicated by the energy requirement, saturation kinetics and a narrow pH profile around pH 6.8. Methylammonium transport (apparent , per g dry weight at 15°C) is competitively inhibited by ammonium (apparent ). The low value and the finding that methylammonium cannot serve as a nitrogen source indicate that ammonium rather than methylammonium is the natural substrate. Uphill transport is driven by a component of the protonmotive force, probably the membrane potential. The transport system is under genetic control; it is partially repressed by amino acids and completely by ammonium. Analysis of mutants suggest that the synthesis of the ammonium transport system is subject to the same ‘nitrogen control’ as nitrogenase and glutamine synthetase. 相似文献
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The kinetics of fructose uptake was determined in perfused rat liver during steady-state fructose elimination. On the basis of the corresponding values of fructose concentration in the affluent and in the effluent medium, and the fructose and ATP concentration in biopsies, the kinetics of membrane transport and intracellular phosphorylation in the intact organ was calculated according to a model system. Carrier-mediated fructose transport has a high (67 mM) and (30 μmoles · min?1 ·g?1). The calculated kinetic constants of the intracellular phosphorylation were compared with values obtained with an acid-treated rat liver high speed supernatant (values given in parentheses). with fructose 1.0 mM (0.7 mM), with ATP 0.54 mM (0.37 mM), 10.3 μmoles · min?1 · g?1 (10.1 μmoles · min?1 · g?1, calculated on the basis of the highest measured rate of fructose uptake correcting the ATP concentration to saturating values). The kinetics of fructose uptake reveals that at Physiological fructose concentrations the membrane transport limits the rate of fructose uptake, thus protecting the liver from severe depletion of adenine nucleotides. 相似文献
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John A. Jacquez 《生物化学与生物物理学报:生物膜》1973,318(3):411-425
The Michaelis-Menten parameters, and of the initial 1-min fluxes of uptake of l-phenylalanine and of α-aminoisobutyric acid were determined for extracellular concentrations of Na+ ranging from 0.5 to 110 mequiv/l for Ehrlich ascites tumor cells. The maximal initial flux, , decreased with decrease in extracellular Na+ for both α-aminoisobutyric acid and phenylalanine but the for α-aminoisobutyric acid increased markedly as the Na+ concentration fell whereas the for phenylalanine decreased. Cycloleucine behaved like phenylalanine.The data provides strong evidence that the Na+-independent flux of phenylalanine is an exchange diffusion flux that can be varied by changing the intracellular level of amino acids such as phenylalanine. For phenylalanine, cyclolcucine, and methionine this exchange diffusion flux appears to be additive with the Na+-dependent initial flux. α-Aminoisobutyric acid also has an exchange diffusion that is Na+-independent but it has a high and is not additive with the Na+-dependent flux. 相似文献
13.
Joseph Y. Cheung David M. Regen Madge E. Schworer Carol F. Whitfield Howard E. Morgan 《生物化学与生物物理学报:生物膜》1977,470(2):212-229
The kinetic parameters of the sugar transport in avian erythrocytes were evaluated under aerobic and anaerobic conditions. In anaerobic cells, transport measurements with methylglucose resulted in a set of similar dissociation-like constants. Thus the Michaelis constants of methylglucose entry and exit, and , were 8 and 7 mM, respectively. The equilibrium exchange constant, , and the counterflow constant, , were 9 and 11 mM, respectively. The activity constant for transport, , defined as , was 4 ml/h per g. This set of kinetic constants was compatible with a symmetrical mobile-carrier model. In contrast, the Michaelis constant for glucose entry, , was 2 mM and less than the counterflow constant, (8 mM). This result could be accounted for by slower movement of the glucose-carrier complex than the free carrier. The activity constant for glucose transport, , was 5 ml/h perg.Under aerobic conditions, two of the dissociation-like constants ( and ) for transport were significantly larger than those obtained in anaerobic cells, but the remaining two ( and ) remained unchanged. The values, for were 8, 26, 20 and 8 mM, respectively. The activity constant, , decreased to 2 ml/h per g. These changes in kinetic constants were consistent with the hypothesis that anoxia accelerated sugar transport by releasing free carrier that was previously sequestered on the inside of the cell membrane. 相似文献
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Phosphate uptake by yeast at pH 7.2 is mediated by two mechanisms, one of which has a of 30 μM and is independent of sodium, and a sodium-dependent mechanism with a of 0.6 μM, both values with respect to monovalent phosphate. The sodium-dependent mechanism has two sites with affinity for Na+, with affinity constants of 0.04 and 29 mM. Also lithium enhances phosphate uptake; the affinity constants for lithium are 0.3 and 36 mM. Other alkali ions do not stimulate phosphate uptake at pH 7.2. Rubidium has no effect on the stimulation of phosphate uptake by sodium.Phosphate and arsenate enhance sodium uptake at pH 7.2. The of this stimulation with regard to monovalent orthophosphate is about equal to that of the sodium-dependent phosphate uptake.The properties of the cation binding sites of the phosphate uptake mechanism and those of the phosphate-dependent cation transport mechanism have been compared. The existence of a separate sodium-phosphate cotransport system is proposed. 相似文献
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G.D. Holman 《生物化学与生物物理学报:生物膜》1980,599(1):202-213
Glucose transport in human erythrocytes is characterized by a marked asymmetry in the and values for entry and for exit. In addition, they show a high and a high for equilibrium exchange but low values for infinite cis and for infinite trans exit and entry. An allosteric pore model has been proposed to account for these characteristics. In this model, substrate-induced conformational changes destabilize the interfaces between protein subunits (the pore gates).Pores doubly occupied from inside destabilize the transport gates and result in high and high transport parameters. This effect is less marked when pores are doubly occupied from outside and therefore transport asymmetry results. 相似文献
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John A. Jacquez 《生物化学与生物物理学报:生物膜》1983,727(2):367-378
Depletion of energy stores of human red cells decreases the maximum transport capacity, , for glucose transport to a value one-third or less of that found in red cells from freshly drawn blood. There is no change in . Hemolysis and resealing of red cells with ATP or ADP reverses the decrease in . The maximum effect occurs at concentrations of ATP in the normal range for red cells, however, there is little effect from ADP concentrations in its normal range in freshly drawn red cells. Hemolysis and resealing with ATP gives an increase in and an increase in differential labeling by photolytic labeling with tritiated cytochalasin B. Most of the activation is lost after a second hemolysis-reseal without ATP but about 25% of the activation remains. 相似文献
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R.M. Johnstone 《生物化学与生物物理学报:生物膜》1978,512(1):199-213
Influx and efflux of glycine have been examined as a function of external and internal Na+ concentrations, respectively, when . With it was found that at comparable external and cellular Na+ levels, the for efflux was larger by an order of magnitude than the value for influx and the for efflux was several times greater than the for influx. For both fluxes the major effect of Na+ was to decrease the value. The observations are consistent with the conclusion that the Na+-dependent transport system is asymmetric per se. Influx and efflux of glycine were increased in a near linear manner by increasing the Na+ concentration from 13 to 100 mM, the half-time for glycine equilibration being a function of the Na+ concentration in absence of an electrochemical potential difference for Na+. In Na+-free media () equilibration of glycine between cells and medium was not achieved after 60 min at 25°C. With , efflux (or uptake) of glycine was not affected by internal (or external) K+ between 20 and 120 mM suggesting that K+ plays no direct role in Na+-dependent transport of glycine in Ehrlich cells. 相似文献
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Bernard Lacour Tilman Drüeke Evelyne Pierandreï Bernadette Nabarra Jean-Louis Funck-Brentano 《生物化学与生物物理学报:生物膜》1981,648(2):151-161
Parathyroid hormone (PTH) and calcitonin exert well known effects on the renal tubule which are thought to involve specific hormone receptors and adenyl cyclase. In the intestine, it is not clear whether the action of PTH and calcitonin is only indirect or also direct, and their mechanisms of action are much less well established. In the present study, possibly direct effects of PTH and calcitonin on Na+ transport in isolated intestinal epithelial cells of rats were investigated. In the presence of bovine PTH (1.2 I.U./ml) in the incubation medium, the Na+ efflux rate constant () of isolated enterocytes was significantly reduced when compared to that in control experiments with the hormone vehicle only. The mean depression of induced by bovine PTH was 26% as compared to the control (100%) and to that induced by ouabain (4.0mM) which was 44%. No depressant effect of bovine PTH on was observed when the isolated enterocytes were incubated with ouabain (4.0 mM). Thus, bovine PTH appeared to inhibit the ouabain-sensitive Na+ pump. When incubating the isolated epithelial cells in an EGTA-containing Ca2+-free medium, bovine PTH lost its capacity to inhibit (). Thus, the presence of extracellular Ca2+ appeared necessary for the inhibitory effect of bovine PTH. In contrast to its effect on , bovine PTH induced no change in net Na+ uptake by isolated enterocytes. Moreover, no significant effect on enterocyte Na+ transport could be demostrated for salmon or porcine calcitonin at two different concentrations in the incubation medium. Neither bovine PTH nor salmon calcitonin induced significant changes in enterocyte cyclic AMP or cyclic GMP concentrations. It was concluded that bovine PTH, but not calcitonin, exerted a direct inhibitory effect on the ouabain-sensitive of isolated rat enterocytes. The effect of bovine PTH occured without measurable activation of the cyclic nucleotide system but needed the presence of Ca2+ in the incubation medium to be operative. 相似文献