共查询到20条相似文献,搜索用时 31 毫秒
1.
Richard J. Naftalin 《The Journal of membrane biology》2010,234(2):75-112
Most membrane transport models are determinate, requiring the transported ligand(s) to bind initially to a vacant site, which
undergoes translation and releases ligand to the alternate side. The carrier reverts to its initial position to complete the
net transport cycle. Ligand affinity may change during translation, but this must be compensated by an equivalent energy change(s)
within the transport cycle. However, any asymmetric cyclic equilibrium deduced on this basis is thermodynamically fallacious.
Determinate cotransport models imply lossless stoichiometric relationships between the complexed cotransported ligands. Independent
ligand leakage apart from the mobile cotransport complex must occur outside the canonical cotransport pathway. In contrast,
stochastic transport models assume independent ligand diffusion through a variably occluded channel(s) containing binding
sites where ligands may undergo bimolecular exchanges. Energy dissipation is intrinsic to all stochastic transport models
and occurs within the primary transport pathway. Frictional interactions within a shared path generate flow coupling between
ligands. The primary driving forces causing transmembrane ligand flows are their electrochemical potential differences between
the external solutions. Demonstrations that ligand exchanges in CLC and neurotransmitter transporters can be multimodal, encompassing
both “channel”-like high and “transporter”-like lower conductance states and have independently regulated import and export
exchange fluxes are major challenges to determinate models but are explicable by transient widening of a close-encounter region
within the channel, leading to decreased coupling and enhanced efflux. 相似文献
2.
Increased Na+-taurine symporter in rat renal brush border membranes: preformed or newly synthesized? 总被引:1,自引:0,他引:1
The renal adaptive response to a varied intake of sulfur amino acids is demonstrated by an increase in the initial rate of Na+-taurine symport (cotransport) by rat renal brush border membrane vesicles (BBMVs) after 8-14 days of a low methionine diet. A high (3%) taurine diet reduces Na+-taurine symport. Fasting for 3 days, which depletes renal tubule cell taurine content, also enhances Na+-taurine symport both initially (15 s) and throughout the overshoot. In this study we examine the possibility that a rapid-onset adaptive response is expressed in BBMV, with the increased Na+-taurine symport reflecting the incorporation of preformed symporter into membranes rather than new synthesis. Rats fed the low methionine diet for 14 days were placed on the high taurine diet for 12-18 h; Na+-taurine symport activity fell by 40%. Fasting for 4 h restored low methionine diet levels of Na+-taurine symport activity (92 pmol.mg protein-1.15 s-1), defining a rapidly induced rise in uptake. Colchicine (0.6 mg) was injected prior to fasting in a group of rats because it blocks the incorporation (import) of preformed symporter into the membrane. Animals injected with colchicine had a pattern of BBMV uptake similar to that found in animals switched to the high taurine diet for 18 h. This agent blocked the rapidly induced rise in uptake. Feeding with the high taurine diet for 4 h caused a fall in uptake of 16.5%; colchicine blocked this reduction in uptake. These results indicate that the nephron can respond rapidly to changes in the intake of amino acids, conserving taurine in periods of nutrient lack and excreting excess taurine within 4 h in periods of surfeit. This rapid response is expressed at the brush border surface. The use of cholchicine indicates that the increase or reduction in Na+-taurine symport activity is due to incorporation (import) of transporter into the BBMV rather than to de novo synthesis. 相似文献
3.
R. Parthasarathy William R. Harvey 《Archives of insect biochemistry and physiology》1995,28(3):237-245
The arginine-specific reagent, phenylglyoxal, decreases the initial rate of lysine/K+ symport (cotransport) as well as maximum lysine accumulation at pH 9.2, by brush border membrane vesicles obtained from the larval midgut of the lepidopteran, Manduca sexta. The symport of a neutral amino acid, leucine, remained unaffected. Following exposure to phenylglyoxal, the apparent dissociation constant for lysine increased by a factor of 2.5 whereas the maximum uptake rate decreased by a factor of 0.4. More than one arginine residue appears to react with phenylglyoxal. Apparently phenylgyoxal reacts preferentially with arginine residues on a symporter that is specific for positively charged lysine. Phenylglyoxal shows promise as a specific covalent label for the identification of a cationic amino acid symporter. © 1995 Wiley-Liss, Inc. 相似文献
4.
Transcellular transport in epithelial cells plays an important role in providing such physiological functions as excretion of cytotoxic substances or reabsorption of metabolites useful for the body life activity. These functions have been shown to be performed by the mechanisms - symport, antiport, ion pumps, and channels - that often function cooperatively. Kinetic models of the substrate transport with the aid of the above mechanisms are widely described in the literature. Much less attention is paid to modeling of cooperative activity of transporters that have different transport mechanisms. In this work we propose a mathematical model for flux coupling of three transporters - the ion pump, symporter, and antiporter as well as of two substrates, one of which (A) can be transported simultaneously by the symport and antiport mechanisms, while the other (B) - only by the latter mechanisms. Analysis of the model has shown that for the pair of substrates (A and B) the flux coupling becomes possible if the following conditions are met: (1) the substrate A flux into the internal cell volume using the symport mechanism is to exceed its antiporter-realized flux in the opposite direction; (2) probability of reorientation from one side of membrane to the other side for the antiporter loaded with the substrate is to be essentially higher than that for empty transporter. The proposed model can be used for comparing efficiency both of excretion and of reabsorption of cell metabolites in representatives of different taxa. 相似文献
5.
Transcellular transport in epithelial cells plays an important role in providing such physiological functions as excretion of cytotoxic substances or reabsorption of metabolites useful for the body life activity. These functions have been shown to be performed by the mechanisms—symport, antiport, ion pumps, and channels—that often function cooperatively. Models for kinetic peculiarities of the substrate transport with the aid of the above mechanisms are widely described in the literature. Much less attention is paid to modeling of cooperative activity of transporters that have different transport mechanisms. In this work we propose a mathematical model for flux coupling of three transporters—the ion pump, symporter, and antiporter as well as of two substrates, one of which (A) can be transported simultaneously by the symport and antiport mechanisms, while the other (B)—only by the latter mechanism. Analysis of the model has shown that for the pair of substrates (A and B) the flux coupling becomes possible if the following conditions are met: (1) the substrate A flux into the internal cell volume using the symport mechanism is to exceed its antiporter-realized flux in the opposite direction; (2) probability of reorientation from one side of membrane to the other side for the antiporter loaded with the substrate is to be essentially higher than that for empty transporter. The proposed model can be used for comparing efficiency both of excretion and of reabsorption of cell metabolites in representatives of different taxa. 相似文献
6.
Cation specificity for sugar substrates of the melibiose carrier in Escherichia coli 总被引:3,自引:0,他引:3
A study has been made of the sugar substrate specificities and the cation specificities of the melibiose transport system of Escherichia coli. The following beta-galactosides were found to be transported: lactose, L-arabinose-beta-D-galactoside, D-fructose-beta-D-galactoside, o- and p-nitrophenyl-beta-D-galactosides. These beta-galactosides were cotransported with Na+ but not with H+. The alpha-galactosides raffinose, melibiose and p-nitrophenyl-alpha-galactoside were transported with either H+ or Na+. Of the monosaccharides tested D-galactose could use either Na+ or H+ for cotransport whereas D-fucose, L-arabinose and D-galactosamine could use only Na+. The sugar specificity requirements for H+ cotransport are therefore more exacting than those for Na+ cotransport. 相似文献
7.
A detailed study of hypertonically stimulated Na-K-2Cl cotransport (NKCC1) in Xenopus laevis oocytes was carried out to better understand the 1 K(+):1 Cl(-) stoichiometry of transport that was previously observed. In this study, we derived the velocity equations for K(+) influx under both rapid equilibrium assumptions and combined equilibrium and steady-state assumptions and demonstrate that the behavior of the equations and curves in Lineweaver-Burke plots are consistent with a model where Cl(-) binds first, followed by Na(+), a second Cl(-), and then K(+). We further demonstrate that stimulation of K(+) movement by K(+) on the trans side is an intrinsic property of a carrier that transports multiple substrates. We also demonstrate that K(+) movement through NKCC1 is strictly dependent upon the presence of external Na(+), even though only a fraction of Na(+) is in fact transported. Finally, we propose that the larger transport of K(+), as compared with Na(+), is a result of the return of partially unloaded carriers, which masks the net 1Na(+):1K(+):2Cl(-) stoichiometry of NKCC1. These data have profound implications for the physiology of Na-K-2Cl cotransport, since transport of K-Cl in some conditions seems to be uncoupled from the transport of Na-Cl. 相似文献
8.
The glutamine/amino acid transporter solubilized from rat renal apical plasma membrane (brush-border membrane) with C12E8 and reconstituted into liposomes has been previously identified as the ASCT2 transporter. The reconstituted transporter catalyses an antiport reaction in which external glutamine and Na+ are cotransported in exchange with internal glutamine (or other amino acids). The glutamine-Na+ cotransport occurred with a 1:1 stoichiometry. The concentration of Na+ did not influence the Km for glutamine and vice versa. Experimental data obtained by a bi-substrate analysis of the glutamine-Na+ cotransport, together with previous report on the glutamine(ex)/glutamine(in) pseudo bi-reactant analysis, indicated that the transporter catalyses a three-substrate transport reaction with a random simultaneous mechanism. The presence of ATP in the internal compartment of the proteoliposomes led to an increase of the Vmax of the transport and to a decrease of the Km of the transporter for external Na+. The reconstituted glutamine/amino acid transporter was inhibited by glutamate; the inhibition was more pronounced at acidic pH. A kinetic analysis revealed that the inhibition was competitive with respect to glutamine. Glutamate was also transported in exchange with glutamine. The external Km of the transporter for glutamate (13.3 mM) was slightly higher than the internal one (8.3 mM). At acidic pH the external but not the internal Km decreased. According with the Km values, glutamate should be transported preferentially from inside to outside in exchange for external glutamine and Na+. 相似文献
9.
The mechanism of pantothenate transport into rabbit renal brush-border membrane vesicles was studied. Under voltage-clamped conditions, an inward NaCl gradient induced the transient accumulation of pantothenate against its concentration gradient, indicating Na+/pantothenate cotransport. K+, Rb+, Li+, NH4+, and choline+ were ineffective in replacing Na+. Pantothenate analogs, D-glucose, and various carboxylic acids did not inhibit Na+-dependent pantothenate transport, suggesting that this system is specific for pantothenate. Kinetic analysis of the Na+-dependent pantothenate uptake revealed a single transport system which obeyed Michaelis-Menten kinetics (Km = 16 microM and Vmax = 6.7 pmol X mg-1 X 10 s-1). Imposition of an inside-negative membrane potential caused net uphill pantothenate accumulation in the presence of Na+ but absence of a Na+ gradient, indicating that Na+/pantothenate cotransport is electrogenic. The relationship between extravesicular Na+ concentration and pantothenate transport measured under voltage-clamped conditions was sigmoidal: a Hill coefficient (napp) of 2 and a [Na+]0.5 of 55 mM were calculated. It is suggested that an anionic pantothenate1- molecule is cotransported with two Na+ to give a net charge of +1. The coupling of pantothenate transport to the Na+ electrochemical gradient may provide an efficient mechanism for reabsorption of pantothenate in the kidney. 相似文献
10.
Mechanism of L-glutamate transport in membrane vesicles from Bacillus stearothermophilus. 总被引:8,自引:5,他引:3
下载免费PDF全文
![点击此处可从《Journal of bacteriology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
In the presence of electrochemical energy, several branched-chain neutral and acidic amino acids were found to accumulate in membrane vesicles of Bacillus stearothermophilus. The membrane vesicles contained a stereo-specific transport system for the acidic amino acids L-glutamate and L-aspartate, which could not translocate their respective amines, L-glutamine and L-asparagine. The transport system was thermostable (Ti = 70 degrees C) and showed highest activities at elevated temperatures (60 to 65 degrees C). The membrane potential or pH gradient could act as the driving force for L-glutamate uptake, which indicated that the transport process of L-glutamate is electrogenic and that protons are involved in the translocation process. The electrogenic character implies that the anionic L-glutamate is cotransported with at least two monovalent cations. To determine the mechanistic stoichiometry of L-glutamate transport and the nature of the cotranslocated cations, the relationship between the components of the proton motive force and the chemical gradient of L-glutamate was investigated at different external pH values in the absence and presence of ionophores. In the presence of either a membrane potential or a pH gradient, the chemical gradient of L-glutamate was equivalent to that specific gradient at different pH values. These results cannot be explained by cotransport of L-glutamate with two protons, assuming thermodynamic equilibrium between the driving force for uptake and the chemical gradient of the substrate. To determine the character of the cotranslocated cations, L-glutamate uptake was monitored with artificial gradients. It was established that either the membrane potential, pH gradient, or chemical gradient of sodium ions could act as the driving force for L-glutamate uptake, which indicated that L-glutamate most likely is cotranslocated in symport with one proton and on sodium ion. 相似文献
11.
Francesca Oppedisano 《生物化学与生物物理学报:生物膜》2007,1768(2):291-298
The glutamine/amino acid transporter solubilized from rat renal apical plasma membrane (brush-border membrane) with C12E8 and reconstituted into liposomes has been previously identified as the ASCT2 transporter. The reconstituted transporter catalyses an antiport reaction in which external glutamine and Na+ are cotransported in exchange with internal glutamine (or other amino acids). The glutamine-Na+ cotransport occurred with a 1:1 stoichiometry. The concentration of Na+ did not influence the Km for glutamine and vice versa. Experimental data obtained by a bi-substrate analysis of the glutamine-Na+ cotransport, together with previous report on the glutamineex/glutaminein pseudo bi-reactant analysis, indicated that the transporter catalyses a three-substrate transport reaction with a random simultaneous mechanism. The presence of ATP in the internal compartment of the proteoliposomes led to an increase of the Vmax of the transport and to a decrease of the Km of the transporter for external Na+. The reconstituted glutamine/amino acid transporter was inhibited by glutamate; the inhibition was more pronounced at acidic pH. A kinetic analysis revealed that the inhibition was competitive with respect to glutamine. Glutamate was also transported in exchange with glutamine. The external Km of the transporter for glutamate (13.3 mM) was slightly higher than the internal one (8.3 mM). At acidic pH the external but not the internal Km decreased. According with the Km values, glutamate should be transported preferentially from inside to outside in exchange for external glutamine and Na+. 相似文献
12.
M L Jennings 《The Journal of membrane biology》1976,28(2-3):187-205
Transient extracellular pH changes accompany the exchange of chloride for sulfate across the erythrocyte membrane. The direction of the extracellular pH change during chloride efflux and sulfate influx depends on experimental conditions. When bicarbonate is present, the extracellular pH drops sharply at the outset of the anion exchange and tends to follow the partial ionic equilibrium described by Wilbrandt (W. Wilbrandt, 1942. Pfluegers Arch. 246:291). When bicarbonate is absent, however, the anion exchange causes the pH to rise, indicating that protons are cotransported with sulfate during chloride-sulfate exchange. The pH rise can be reversed by the addition of HCO(-3) (4 muM) or 2,4-dinitrophenol (90 muM). This demonstrates that the proton-sulfate cotransport can drive proton transport uphill. The stoichiometry of the transport is that one chloride exchanges for one sulfate plus one proton. These results support the titratable carrier model proposed by Gunn (Gunn, R.B. 1972, In: Oxygen Affinity of Hemoglobin and Red Cell Acid-Base Status. M. Rorth and P. Astrup, editors. p. 823. Munksgaard, Copenhagen) for erythrocyte membrane anion exchange. 相似文献
13.
In order to clarify contradictory reports on the electrogenicity of sodium/L-glutamate cotransport, this cotransport was studied using brush-border membrane vesicles isolated from rabbit renal cortex. Beforehand, the claim that the symport of L-glutamate with Na+ is linked to simultaneous antiport with K+ has been confirmed by the demonstration that equilibrium exchange of L-glutamate is inhibited by potassium. Concerning the electrogenicity of the system, the following results are reported: net uptake of sodium-dependent L-glutamate uptake was stimulated when the transmembranal electrical potential difference was increased by replacing a sodium sulfate gradient by a sodium nitrate gradient. At 100 mM Na+ the 'relative electrogenicity' of the initial uptake in the presence of intravesicular potassium was 2-times higher than in its absence. At a sodium concentration of 20 mM, when overall uptake was reduced, the relative electrogenicity in the presence of K+ was even 3-fold higher than in K+-free media. The relative electrogenicity of sodium/D-glucose cotransport measured under the same experimental conditions was not affected by K+. These results are discussed in terms of a model where the apparent electrogenicity of a cotransport system is dependent on the extent to which the charge translocating step is rate limiting ('rate limitancy'). It is proposed that potassium antiport, while decreasing charge stoichiometry of Na+/glutamate transport, increases the relative rate limitancy of the transport step translocating three cations (probably two Na+, one H+) together with one glutamate. Thereby the positive electrogenicity of glutamate uptake increases, in complete contrast to what would be expected from simple considerations of charge stoichiometry. 相似文献
14.
Previous work with L-[3H]glutamate transport by lobster (Homarus americanus) hepatopancreatic brush border membrane vesicles (BBMV) indicated that the transport of this amino acid was stimulated by the presence of both Na+ and Cl- ions in the external medium, however, the specific catalytic or energetic role of each monovalent ion in amino acid transfer was not established (Ahearn and Clay (1987) J. Exp. Biol. 130, 175-191). The present study employs a variety of experimental treatments with this membrane preparation to clarify the nature of the ion dependency in the cotransport process. A zero-trans time course experiment using inwardly-directed transmembrane Na+ or Cl- gradients led to similar transient accumulations of the amino acid above equilibrium values in the presence of equilibrated concentrations of the respective counterions. The uptake overshoots observed in the presence of single ion gradients were significantly increased when gradients of both Na+ and Cl- were used simultaneously. When vesicles were pre-equilibrated with L-[3H]glutamate and either of the monovalent ions, an inwardly-directed gradient of each counterion led to the transient accumulation of additional labelled amino acid above its equilibrium concentration, indicating that either ion gradient was capable of energizing the net flow of L-glutamate. A cotransport stoichiometry of 1 Na+/1 Cl-/1 L-glutamate was established using the Static Head analysis where a balance of ion and amino acid driving forces were attained with a 7:1 Na+ or Cl- gradient (o greater than i) against a 7:1 L-glutamate gradient (i greater than o). 相似文献
15.
16.
Three parallel pathways of l-lactate transport across the membrane of human red blood cells can be discriminated: (a) by nonionic diffusion; (b) via the band 3 anion exchange protein; and (c) via a specific monocarboxylate carrier system. Influx of lactate via the latter system leads to alkalinization of the medium, suggesting lactate-proton symport. Kinetic analysis of initial lactate influx via the monocarboxylate carrier indicates a symport system with ordered binding of the two ligands, in the sense that a proton binds first to the translocator, followed by lactate binding to the protonated carrier. The influence of varying trans-pH under conditions of net (zero-trans) flux with constant cis-pH indicates that the monocarboxylate translocator should be considered as a mobile carrier, with the ligand-binding sites exposed alternately to the outside and the inside of the membrane. 相似文献
17.
Kinetic analysis of L-lactate transport in human erythrocytes via the monocarboxylate-specific carrier system 总被引:2,自引:0,他引:2
Three parallel pathways of L-lactate transport across the membrane of human red blood cells can be discriminated: (a) by nonionic diffusion; (b) via the band 3 anion exchange protein; and (c) via a specific monocarboxylate carrier system. Influx of lactate via the latter system leads to alkalinization of the medium, suggesting lactate-proton symport. Kinetic analysis of initial lactate influx via the monocarboxylate carrier indicates a symport system with ordered binding of the two ligands, in the sense that a proton binds first to the translocator, followed by lactate binding to the protonated carrier. The influence of varying trans-pH under conditions of net (zero-trans) flux with constant cis-pH indicates that the monocarboxylate translocator should be considered as a mobile carrier, with the ligand-binding sites exposed alternatively to the outside and the inside of the membrane. 相似文献
18.
A detailed kinetic study of K:Cl cotransport in hyposmotically swollen low K sheep red blood cells was carried out to characterize the nature of the outwardly poised carrier. The kinetic parameters were determined from the rate of K efflux and influx under zero-K-trans conditions in red cells with cellular K altered by the nystatin method and with different extracellular K or Rb concentrations. Although apparent affinities for efflux and influx were quite similar, the maximal velocity for K efflux was approximately two times greater than for influx. Furthermore, at thermodynamic equilibrium (i.e., when the ion product of K and Cl within the cell was equal to that outside) a temperature-dependent net K efflux was observed, approaching zero only when the external product reached approximately two times the internal product. The binding order of the ions to the transporter was asymmetric, being ordered outside (Cl binding first, followed by K) and random inside. K efflux but not influx was trans-inhibited by KCl. Trans inhibition of K efflux was used to verify the order of binding outside: trans inhibition by external Cl occurred in the absence of external K, but not vice versa. Thus K:Cl cotransport is kinetically asymmetric in hyposmotically swollen low K sheep red cells. 相似文献
19.
A member of the sugar transporter family, Stl1p is the glycerol/H+ symporter in Saccharomyces cerevisiae
下载免费PDF全文
![点击此处可从《Molecular biology of the cell》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Ferreira C van Voorst F Martins A Neves L Oliveira R Kielland-Brandt MC Lucas C Brandt A 《Molecular biology of the cell》2005,16(4):2068-2076
Glycerol and other polyols are used as osmoprotectants by many organisms. Several yeasts and other fungi can take up glycerol by proton symport. To identify genes involved in active glycerol uptake in Saccharomyces cerevisiae we screened a deletion mutant collection comprising 321 genes encoding proteins with 6 or more predicted transmembrane domains for impaired growth on glycerol medium. Deletion of STL1, which encodes a member of the sugar transporter family, eliminates active glycerol transport. Stl1p is present in the plasma membrane in S. cerevisiae during conditions where glycerol symport is functional. Both the Stl1 protein and the active glycerol transport are subject to glucose-induced inactivation, following identical patterns. Furthermore, the Stl1 protein and the glycerol symporter activity are strongly but transiently induced when cells are subjected to osmotic shock. STL1 was heterologously expressed in Schizosaccharomyces pombe, a yeast that does not contain its own active glycerol transport system. In S. pombe, STL1 conferred the ability to take up glycerol against a concentration gradient in a proton motive force-dependent manner. We conclude that the glycerol proton symporter in S. cerevisiae is encoded by STL1. 相似文献
20.
The relationship between Na/K/2Cl cotransport activation in duck erythrocytes and binding of the diuretic [3H]bumetanide to isolated membranes from stimulated cells has been assessed. Cotransport was activated by either cAMP-dependent (norepinephrine) or -independent (fluoride, hypertonicity) pathways. Membranes isolated from unstimulated cells possessed no specific bumetanide binding. In the presence of norepinephrine, cotransport and saturable binding rose in parallel, reaching a maximum after 5-7 min. In membranes from maximally stimulated cells the K1/2 and Bmax for bumetanide binding were 100 nM and 1.7 pmol/mg protein, respectively. The diuretic binding properties of these membranes were characteristic of interactions of ligands with the Na/K/2Cl cotransporter: specific binding required the presence of all three cotransported ions (Na, K, and Cl), and the rank order of potency for diuretic competition with bumetanide for binding sites was benzmetanide greater than bumetanide greater than furosemide. The appearance of specific bumetanide binding was also seen in membranes from erythrocytes activated by non-cAMP-dependent stimuli, with an excellent temporal correlation between cotransport activation and diuretic binding. On removal of all stimuli both cotransport and bumetanide binding declined in parallel. Duck erythrocytes treated with norepinephrine in a solution containing 15 mM K+ swell to a new stable cell volume after 60 min, during which time cotransport becomes inoperative. Bumetanide binding to both whole cells and isolated membranes paralleled the decline in cotransport activity. It is concluded that bumetanide binding to isolated membranes faithfully reflects the state of activation of the Na/K/2Cl cotransporter in intact cells under a variety of conditions. 相似文献