Cation-Coupled Bicarbonate Transporters

PubMed Central

Aalkjaer, Christian; Boedtkjer, Ebbe; Choi, Inyeong; Lee, Soojung

2016-01-01

Cation-coupled HCO3− transport was initially identified in the mid-1970s when pioneering studies showed that acid extrusion from cells is stimulated by CO2/HCO3− and associated with Na+ and Cl− movement. The first Na+-coupled bicarbonate transporter (NCBT) was expression-cloned in the late 1990s. There are currently five mammalian NCBTs in the SLC4-family: the electrogenic Na,HCO3-cotransporters NBCe1 and NBCe2 (SLC4A4 and SLC4A5 gene products); the electroneutral Na,HCO3-cotransporter NBCn1 (SLC4A7 gene product); the Na+-driven Cl,HCO3-exchanger NDCBE (SLC4A8 gene product); and NBCn2/NCBE (SLC4A10 gene product), which has been characterized as an electroneutral Na,HCO3-cotransporter or a Na+-driven Cl,HCO3-exchanger. Despite the similarity in amino acid sequence and predicted structure among the NCBTs of the SLC4-family, they exhibit distinct differences in ion dependency, transport function, pharmacological properties, and interactions with other proteins. In epithelia, NCBTs are involved in transcellular movement of acid-base equivalents and intracellular pH control. In nonepithelial tissues, NCBTs contribute to intracellular pH regulation; and hence, they are crucial for diverse tissue functions including neuronal discharge, sensory neuron development, performance of the heart, and vascular tone regulation. The function and expression levels of the NCBTs are generally sensitive to intracellular and systemic pH. Animal models have revealed pathophysiological roles of the transporters in disease states including metabolic acidosis, hypertension, visual defects, and epileptic seizures. Studies are being conducted to understand the physiological consequences of genetic polymorphisms in the SLC4-members, which are associated with cancer, hypertension, and drug addiction. Here, we describe the current knowledge regarding the function, structure, and regulation of the mammalian cation-coupled HCO3− transporters of the SLC4-family. PMID:25428855

Role of hexose transport in control of glycolytic flux in Saccharomyces cerevisiae.

PubMed

Elbing, Karin; Larsson, Christer; Bill, Roslyn M; Albers, Eva; Snoep, Jacky L; Boles, Eckhard; Hohmann, Stefan; Gustafsson, Lena

2004-09-01

The yeast Saccharomyces cerevisiae predominantly ferments glucose to ethanol at high external glucose concentrations, irrespective of the presence of oxygen. In contrast, at low external glucose concentrations and in the presence of oxygen, as in a glucose-limited chemostat, no ethanol is produced. The importance of the external glucose concentration suggests a central role for the affinity and maximal transport rates of yeast's glucose transporters in the control of ethanol production. Here we present a series of strains producing functional chimeras between the hexose transporters Hxt1 and Hxt7, each of which has distinct glucose transport characteristics. The strains display a range of decreasing glycolytic rates resulting in a proportional decrease in ethanol production. Using these strains, we show for the first time that at high glucose levels, the glucose uptake capacity of wild-type S. cerevisiae does not control glycolytic flux during exponential batch growth. In contrast, our chimeric Hxt transporters control the rate of glycolysis to a high degree. Strains whose glucose uptake is mediated by these chimeric transporters will undoubtedly provide a powerful tool with which to examine in detail the mechanism underlying the switch between fermentation and respiration in S. cerevisiae and will provide new tools for the control of industrial fermentations.

Role of Hexose Transport in Control of Glycolytic Flux in Saccharomyces cerevisiae

PubMed Central

Elbing, Karin; Larsson, Christer; Bill, Roslyn M.; Albers, Eva; Snoep, Jacky L.; Boles, Eckhard; Hohmann, Stefan; Gustafsson, Lena

2004-01-01

The yeast Saccharomyces cerevisiae predominantly ferments glucose to ethanol at high external glucose concentrations, irrespective of the presence of oxygen. In contrast, at low external glucose concentrations and in the presence of oxygen, as in a glucose-limited chemostat, no ethanol is produced. The importance of the external glucose concentration suggests a central role for the affinity and maximal transport rates of yeast's glucose transporters in the control of ethanol production. Here we present a series of strains producing functional chimeras between the hexose transporters Hxt1 and Hxt7, each of which has distinct glucose transport characteristics. The strains display a range of decreasing glycolytic rates resulting in a proportional decrease in ethanol production. Using these strains, we show for the first time that at high glucose levels, the glucose uptake capacity of wild-type S. cerevisiae does not control glycolytic flux during exponential batch growth. In contrast, our chimeric Hxt transporters control the rate of glycolysis to a high degree. Strains whose glucose uptake is mediated by these chimeric transporters will undoubtedly provide a powerful tool with which to examine in detail the mechanism underlying the switch between fermentation and respiration in S. cerevisiae and will provide new tools for the control of industrial fermentations. PMID:15345416

Allelic variants of hexose transporter Hxt3p and hexokinases Hxk1p/Hxk2p in strains of Saccharomyces cerevisiae and interspecies hybrids.

PubMed

Zuchowska, Magdalena; Jaenicke, Elmar; König, Helmut; Claus, Harald

2015-11-01

The transport of sugars across the plasma membrane is a critical step in the utilization of glucose and fructose by Saccharomyces cerevisiae during must fermentations. Variations in the molecular structure of hexose transporters and kinases may affect the ability of wine yeast strains to finish sugar fermentation, even under stressful wine conditions. In this context, we sequenced and compared genes encoding the hexose transporter Hxt3p and the kinases Hxk1p/Hxk2p of Saccharomyces strains and interspecies hybrids with different industrial usages and regional backgrounds. The Hxt3p primary structure varied in a small set of amino acids, which characterized robust yeast strains used for the production of sparkling wine or to restart stuck fermentations. In addition, interspecies hybrid strains, previously isolated at the end of spontaneous fermentations, revealed a common amino acid signature. The location and potential influence of the amino acids exchanges is discussed by means of a first modelled Hxt3p structure. In comparison, hexokinase genes were more conserved in different Saccharomyces strains and hybrids. Thus, molecular variants of the hexose carrier Hxt3p, but not of kinases, correlate with different fermentation performances of yeast. Copyright © 2015 John Wiley & Sons, Ltd.

Gastric emptying of hexose sugars: role of osmolality, molecular structure and the CCK₁ receptor.

PubMed

Little, T J; Gopinath, A; Patel, E; McGlone, A; Lassman, D J; D'Amato, M; McLaughlin, J T; Thompson, D G

2010-11-01

It is widely reported that hexose sugars slow gastric emptying (GE) via osmoreceptor stimulation but this remains uncertain. We evaluated the effects of a panel of hexoses of differing molecular structure, assessing the effects of osmolality, intra-individual reproducibility and the role of the CCK(1) receptor, in the regulation of GE by hexoses. Thirty one healthy non-obese male and female subjects were studied in a series of protocols, using a (13) C-acetate breath test to evaluate GE of varying concentrations of glucose, galactose, fructose and tagatose, with water, NaCl and lactulose as controls. GE was further evaluated following the administration of a CCK(1) receptor antagonist. Three subjects underwent repeated studies to evaluate intra-individual reproducibility. At 250 mOsmol, a hexose-specific effect was apparent: tagatose slowed GE more potently than water, glucose and fructose (P < 0.05). Fructose (P < 0.05) also slowed GE, but with substantial inter-, but not intra-, individual differences. As osmolality increased further the hexose-specific differences were lost. At 500 mOsmol, all hexoses slowed GE compared with water (P < 0.05), whereas lactulose and saline did not. The slowing of GE by hexose sugars appeared to be CCK(1) receptor-dependent. The effects of hexose sugars on GE appear related to their molecular structure rather than osmolality per se, and are, at least in part, CCK(1) receptor-dependent. © 2010 Blackwell Publishing Ltd.

KCNQ1, KCNE2, and Na+-Coupled Solute Transporters Form Reciprocally Regulating Complexes that Affect Neuronal Excitability

PubMed Central

Abbott, Geoffrey W.; Tai, Kwok-Keung; Neverisky, Daniel; Hansler, Alex; Hu, Zhaoyang; Roepke, Torsten K.; Lerner, Daniel J.; Chen, Qiuying; Liu, Li; Zupan, Bojana; Toth, Miklos; Haynes, Robin; Huang, Xiaoping; Demirbas, Didem; Buccafusca, Roberto; Gross, Steven S.; Kanda, Vikram A.; Berry, Gerard T.

2014-01-01

Na+-coupled solute transport is crucial for the uptake of nutrients and metabolic precursors, such as myo-inositol, an important osmolyte and precursor for various cell signaling molecules. Here, we found that various solute transporters and potassium channel subunits formed complexes and reciprocally regulated each other in vitro and in vivo. Global metabolite profiling revealed that mice lacking KCNE2, a K+ channel β subunit, showed a reduction in the myo-inositol concentration in cerebrospinal fluid (CSF) but not in serum. Increased behavorial responsiveness to stress and seizure susceptibility in Kcne2−/− mice were alleviated by injections of myo-inositol. Suspecting a defect in myo-inositol transport, we found that KCNE2 and KCNQ1, a voltage-gated potassium channel α subunit, colocalized and coimmunoprecipitated with SMIT1, a Na+-coupled myo-inositol transporter, in the choroid plexus epithelium. Heterologous coexpression demonstrated that myo-inositol transport by SMIT1 was augmented by coexpression of KCNQ1 but inhibited by coexpression of both KCNQ1 and KCNE2, which form a constitutively active, heteromeric K+ channel. SMIT1 and the related transporter SMIT2 were also inhibited by a constitutively active mutant form of KCNQ1. The activity of KCNQ1 and KCNQ1-KCNE2 were augmented by SMIT1 and the glucose transporter SGLT1, but suppressed by SMIT2. Channel-transporter signaling complexes may be a widespread mechanism to facilitate solute transport and electrochemical crosstalk. PMID:24595108

KCNQ1, KCNE2, and Na+-coupled solute transporters form reciprocally regulating complexes that affect neuronal excitability.

PubMed

Abbott, Geoffrey W; Tai, Kwok-Keung; Neverisky, Daniel L; Hansler, Alex; Hu, Zhaoyang; Roepke, Torsten K; Lerner, Daniel J; Chen, Qiuying; Liu, Li; Zupan, Bojana; Toth, Miklos; Haynes, Robin; Huang, Xiaoping; Demirbas, Didem; Buccafusca, Roberto; Gross, Steven S; Kanda, Vikram A; Berry, Gerard T

2014-03-04

Na(+)-coupled solute transport is crucial for the uptake of nutrients and metabolic precursors, such as myo-inositol, an important osmolyte and precursor for various cell signaling molecules. We found that various solute transporters and potassium channel subunits formed complexes and reciprocally regulated each other in vitro and in vivo. Global metabolite profiling revealed that mice lacking KCNE2, a K(+) channel β subunit, showed a reduction in myo-inositol concentration in cerebrospinal fluid (CSF) but not in serum. Increased behavioral responsiveness to stress and seizure susceptibility in Kcne2(-/-) mice were alleviated by injections of myo-inositol. Suspecting a defect in myo-inositol transport, we found that KCNE2 and KCNQ1, a voltage-gated potassium channel α subunit, colocalized and coimmunoprecipitated with SMIT1, a Na(+)-coupled myo-inositol transporter, in the choroid plexus epithelium. Heterologous coexpression demonstrated that myo-inositol transport by SMIT1 was augmented by coexpression of KCNQ1 but was inhibited by coexpression of both KCNQ1 and KCNE2, which form a constitutively active, heteromeric K(+) channel. SMIT1 and the related transporter SMIT2 were also inhibited by a constitutively active mutant form of KCNQ1. The activities of KCNQ1 and KCNQ1-KCNE2 were augmented by SMIT1 and the glucose transporter SGLT1 but were suppressed by SMIT2. Channel-transporter signaling complexes may be a widespread mechanism to facilitate solute transport and electrochemical crosstalk.

Multilevel regulation of an α-arrestin by glucose depletion controls hexose transporter endocytosis.

PubMed

Hovsepian, Junie; Defenouillère, Quentin; Albanèse, Véronique; Váchová, Libuše; Garcia, Camille; Palková, Zdena; Léon, Sébastien

2017-06-05

Nutrient availability controls the landscape of nutrient transporters present at the plasma membrane, notably by regulating their ubiquitylation and subsequent endocytosis. In yeast, this involves the Nedd4 ubiquitin ligase Rsp5 and arrestin-related trafficking adaptors (ARTs). ARTs are targeted by signaling pathways and warrant that cargo ubiquitylation and endocytosis appropriately respond to nutritional inputs. Here, we show that glucose deprivation regulates the ART protein Csr2/Art8 at multiple levels to trigger high-affinity glucose transporter endocytosis. Csr2 is transcriptionally induced in these conditions through the AMPK orthologue Snf1 and downstream transcriptional repressors. Upon synthesis, Csr2 becomes activated by ubiquitylation. In contrast, glucose replenishment induces CSR2 transcriptional shutdown and switches Csr2 to an inactive, deubiquitylated form. This glucose-induced deubiquitylation of Csr2 correlates with its phospho-dependent association with 14-3-3 proteins and involves protein kinase A. Thus, two glucose signaling pathways converge onto Csr2 to regulate hexose transporter endocytosis by glucose availability. These data illustrate novel mechanisms by which nutrients modulate ART activity and endocytosis. © 2017 Hovsepian et al.

Multilevel regulation of an α-arrestin by glucose depletion controls hexose transporter endocytosis

PubMed Central

Hovsepian, Junie; Váchová, Libuše; Garcia, Camille; Palková, Zdena

2017-01-01

Nutrient availability controls the landscape of nutrient transporters present at the plasma membrane, notably by regulating their ubiquitylation and subsequent endocytosis. In yeast, this involves the Nedd4 ubiquitin ligase Rsp5 and arrestin-related trafficking adaptors (ARTs). ARTs are targeted by signaling pathways and warrant that cargo ubiquitylation and endocytosis appropriately respond to nutritional inputs. Here, we show that glucose deprivation regulates the ART protein Csr2/Art8 at multiple levels to trigger high-affinity glucose transporter endocytosis. Csr2 is transcriptionally induced in these conditions through the AMPK orthologue Snf1 and downstream transcriptional repressors. Upon synthesis, Csr2 becomes activated by ubiquitylation. In contrast, glucose replenishment induces CSR2 transcriptional shutdown and switches Csr2 to an inactive, deubiquitylated form. This glucose-induced deubiquitylation of Csr2 correlates with its phospho-dependent association with 14-3-3 proteins and involves protein kinase A. Thus, two glucose signaling pathways converge onto Csr2 to regulate hexose transporter endocytosis by glucose availability. These data illustrate novel mechanisms by which nutrients modulate ART activity and endocytosis. PMID:28468835

Use of a Selective Inhibitor To Define the Chemotherapeutic Potential of the Plasmodial Hexose Transporter in Different Stages of the Parasite's Life Cycle▿

PubMed Central

Slavic, Ksenija; Delves, Michael J.; Prudêncio, Miguel; Talman, Arthur M.; Straschil, Ursula; Derbyshire, Elvira T.; Xu, Zhengyao; Sinden, Robert E.; Mota, Maria M.; Morin, Christophe; Tewari, Rita; Krishna, Sanjeev; Staines, Henry M.

2011-01-01

During blood infection, malarial parasites use d-glucose as their main energy source. The Plasmodium falciparum hexose transporter (PfHT), which mediates the uptake of d-glucose into parasites, is essential for survival of asexual blood-stage parasites. Recently, genetic studies in the rodent malaria model, Plasmodium berghei, found that the orthologous hexose transporter (PbHT) is expressed throughout the parasite's development within the mosquito vector, in addition to being essential during intraerythrocytic development. Here, using a d-glucose-derived specific inhibitor of plasmodial hexose transporters, compound 3361, we have investigated the importance of d-glucose uptake during liver and transmission stages of P. berghei. Initially, we confirmed the expression of PbHT during liver stage development, using a green fluorescent protein (GFP) tagging strategy. Compound 3361 inhibited liver-stage parasite development, with a 50% inhibitory concentration (IC50) of 11 μM. This process was insensitive to the external d-glucose concentration. In addition, compound 3361 inhibited ookinete development and microgametogenesis, with IC50s in the region of 250 μM (the latter in a d-glucose-sensitive manner). Consistent with our findings for the effect of compound 3361 on vector parasite stages, 1 mM compound 3361 demonstrated transmission blocking activity. These data indicate that novel chemotherapeutic interventions that target PfHT may be active against liver and, to a lesser extent, transmission stages, in addition to blood stages. PMID:21402842

Use of a selective inhibitor to define the chemotherapeutic potential of the plasmodial hexose transporter in different stages of the parasite's life cycle.

PubMed

Slavic, Ksenija; Delves, Michael J; Prudêncio, Miguel; Talman, Arthur M; Straschil, Ursula; Derbyshire, Elvira T; Xu, Zhengyao; Sinden, Robert E; Mota, Maria M; Morin, Christophe; Tewari, Rita; Krishna, Sanjeev; Staines, Henry M

2011-06-01

During blood infection, malarial parasites use D-glucose as their main energy source. The Plasmodium falciparum hexose transporter (PfHT), which mediates the uptake of D-glucose into parasites, is essential for survival of asexual blood-stage parasites. Recently, genetic studies in the rodent malaria model, Plasmodium berghei, found that the orthologous hexose transporter (PbHT) is expressed throughout the parasite's development within the mosquito vector, in addition to being essential during intraerythrocytic development. Here, using a D-glucose-derived specific inhibitor of plasmodial hexose transporters, compound 3361, we have investigated the importance of D-glucose uptake during liver and transmission stages of P. berghei. Initially, we confirmed the expression of PbHT during liver stage development, using a green fluorescent protein (GFP) tagging strategy. Compound 3361 inhibited liver-stage parasite development, with a 50% inhibitory concentration (IC₅₀) of 11 μM. This process was insensitive to the external D-glucose concentration. In addition, compound 3361 inhibited ookinete development and microgametogenesis, with IC₅₀s in the region of 250 μM (the latter in a D-glucose-sensitive manner). Consistent with our findings for the effect of compound 3361 on vector parasite stages, 1 mM compound 3361 demonstrated transmission blocking activity. These data indicate that novel chemotherapeutic interventions that target PfHT may be active against liver and, to a lesser extent, transmission stages, in addition to blood stages.

Identification of a 3rd Na+ Binding Site of the Glycine Transporter, GlyT2.

PubMed

Subramanian, Nandhitha; Scopelitti, Amanda J; Carland, Jane E; Ryan, Renae M; O'Mara, Megan L; Vandenberg, Robert J

2016-01-01

The Na+/Cl- dependent glycine transporters GlyT1 and GlyT2 regulate synaptic glycine concentrations. Glycine transport by GlyT2 is coupled to the co-transport of three Na+ ions, whereas transport by GlyT1 is coupled to the co-transport of only two Na+ ions. These differences in ion-flux coupling determine their respective concentrating capacities and have a direct bearing on their functional roles in synaptic transmission. The crystal structures of the closely related bacterial Na+-dependent leucine transporter, LeuTAa, and the Drosophila dopamine transporter, dDAT, have allowed prediction of two Na+ binding sites in GlyT2, but the physical location of the third Na+ site in GlyT2 is unknown. A bacterial betaine transporter, BetP, has also been crystallized and shows structural similarity to LeuTAa. Although betaine transport by BetP is coupled to the co-transport of two Na+ ions, the first Na+ site is not conserved between BetP and LeuTAa, the so called Na1' site. We hypothesized that the third Na+ binding site (Na3 site) of GlyT2 corresponds to the BetP Na1' binding site. To identify the Na3 binding site of GlyT2, we performed molecular dynamics (MD) simulations. Surprisingly, a Na+ placed at the location consistent with the Na1' site of BetP spontaneously dissociated from its initial location and bound instead to a novel Na3 site. Using a combination of MD simulations of a comparative model of GlyT2 together with an analysis of the functional properties of wild type and mutant GlyTs we have identified an electrostatically favorable novel third Na+ binding site in GlyT2 formed by Trp263 and Met276 in TM3, Ala481 in TM6 and Glu648 in TM10.

Identity of SMCT1 (SLC5A8) as a neuron-specific Na+-coupled transporter for active uptake of L-lactate and ketone bodies in the brain.

PubMed

Martin, Pamela M; Gopal, Elangovan; Ananth, Sudha; Zhuang, Lina; Itagaki, Shiro; Prasad, Balakrishna M; Smith, Sylvia B; Prasad, Puttur D; Ganapathy, Vadivel

2006-07-01

SMCT1 is a sodium-coupled (Na(+)-coupled) transporter for l-lactate and short-chain fatty acids. Here, we show that the ketone bodies, beta-d-hydroxybutyrate and acetoacetate, and the branched-chain ketoacid, alpha-ketoisocaproate, are also substrates for the transporter. The transport of these compounds via human SMCT1 is Na(+)-coupled and electrogenic. The Michaelis constant is 1.4 +/- 0.1 mm for beta-d-hydroxybutyrate, 0.21 +/- 0.04 mm for acetoacetate and 0.21 +/- 0.03 mm for alpha-ketoisocaproate. The Na(+) : substrate stoichiometry is 2 : 1. As l-lactate and ketone bodies constitute primary energy substrates for neurons, we investigated the expression pattern of this transporter in the brain. In situ hybridization studies demonstrate widespread expression of SMCT1 mRNA in mouse brain. Immunofluorescence analysis shows that SMCT1 protein is expressed exclusively in neurons. SMCT1 protein co-localizes with MCT2, a neuron-specific Na(+)-independent monocarboxylate transporter. In contrast, there was no overlap of signals for SMCT1 and MCT1, the latter being expressed only in non-neuronal cells. We also demonstrate the neuron-specific expression of SMCT1 in mixed cultures of rat cortical neurons and astrocytes. This represents the first report of an Na(+)-coupled transport system for a major group of energy substrates in neurons. These findings suggest that SMCT1 may play a critical role in the entry of l-lactate and ketone bodies into neurons by a process driven by an electrochemical Na(+) gradient and hence, contribute to the maintenance of the energy status and function of neurons.

Evaluation of the electrophysiological consequences of GABA removal from the synaptic cleft by Na+ ion transport-coupled neuronal uptake.

PubMed

Cupello, A; Hydén, H

1985-12-09

The pre- and postsynaptic electrophysiological consequences of a carrier-mediated, Na+ ion transport-coupled removal of gamma-aminobutyric acid (GABA) from the relevant synaptic clefts are discussed. Assuming for the GABA internalization process a stoichiometry like GABAo + 3NA+o + K+i in equilibrium GABAi + 3Na+i + K+o and a synaptic cleft GABA maximal concentration of 100 microM we calculated the presynaptic depolarization associated with GABA removal between 11.5 and 38.2 mV. At the postsynaptic level the effect appears to be less marked.

Identification of a 3rd Na+ Binding Site of the Glycine Transporter, GlyT2

PubMed Central

Subramanian, Nandhitha; Scopelitti, Amanda J.; Carland, Jane E.; Ryan, Renae M.; O’Mara, Megan L.; Vandenberg, Robert J.

2016-01-01

The Na+/Cl- dependent glycine transporters GlyT1 and GlyT2 regulate synaptic glycine concentrations. Glycine transport by GlyT2 is coupled to the co-transport of three Na+ ions, whereas transport by GlyT1 is coupled to the co-transport of only two Na+ ions. These differences in ion-flux coupling determine their respective concentrating capacities and have a direct bearing on their functional roles in synaptic transmission. The crystal structures of the closely related bacterial Na+-dependent leucine transporter, LeuTAa, and the Drosophila dopamine transporter, dDAT, have allowed prediction of two Na+ binding sites in GlyT2, but the physical location of the third Na+ site in GlyT2 is unknown. A bacterial betaine transporter, BetP, has also been crystallized and shows structural similarity to LeuTAa. Although betaine transport by BetP is coupled to the co-transport of two Na+ ions, the first Na+ site is not conserved between BetP and LeuTAa, the so called Na1' site. We hypothesized that the third Na+ binding site (Na3 site) of GlyT2 corresponds to the BetP Na1' binding site. To identify the Na3 binding site of GlyT2, we performed molecular dynamics (MD) simulations. Surprisingly, a Na+ placed at the location consistent with the Na1' site of BetP spontaneously dissociated from its initial location and bound instead to a novel Na3 site. Using a combination of MD simulations of a comparative model of GlyT2 together with an analysis of the functional properties of wild type and mutant GlyTs we have identified an electrostatically favorable novel third Na+ binding site in GlyT2 formed by Trp263 and Met276 in TM3, Ala481 in TM6 and Glu648 in TM10. PMID:27337045

Histidine residues in the Na+-coupled ascorbic acid transporter-2 (SVCT2) are central regulators of SVCT2 function, modulating pH sensitivity, transporter kinetics, Na+ cooperativity, conformational stability, and subcellular localization.

PubMed

Ormazabal, Valeska; Zuñiga, Felipe A; Escobar, Elizabeth; Aylwin, Carlos; Salas-Burgos, Alexis; Godoy, Alejandro; Reyes, Alejandro M; Vera, Juan Carlos; Rivas, Coralia I

2010-11-19

Na(+)-coupled ascorbic acid transporter-2 (SVCT2) activity is impaired at acid pH, but little is known about the molecular determinants that define the transporter pH sensitivity. SVCT2 contains six histidine residues in its primary sequence, three of which are exofacial in the transporter secondary structure model. We used site-directed mutagenesis and treatment with diethylpyrocarbonate to identify histidine residues responsible for SVCT2 pH sensitivity. We conclude that five histidine residues, His(109), His(203), His(206), His(269), and His(413), are central regulators of SVCT2 function, participating to different degrees in modulating pH sensitivity, transporter kinetics, Na(+) cooperativity, conformational stability, and subcellular localization. Our results are compatible with a model in which (i) a single exofacial histidine residue, His(413), localized in the exofacial loop IV that connects transmembrane helices VII-VIII defines the pH sensitivity of SVCT2 through a mechanism involving a marked attenuation of the activation by Na(+) and loss of Na(+) cooperativity, which leads to a decreased V(max) without altering the transport K(m); (ii) exofacial histidine residues His(203), His(206), and His(413) may be involved in maintaining a functional interaction between exofacial loops II and IV and influence the general folding of the transporter; (iii) histidines 203, 206, 269, and 413 affect the transporter kinetics by modulating the apparent transport K(m); and (iv) histidine 109, localized at the center of transmembrane helix I, might be fundamental for the interaction of SVCT2 with the transported substrate ascorbic acid. Thus, histidine residues are central regulators of SVCT2 function.

Substrate-bound outward-open state of the betaine transporter BetP provides insights into Na+ coupling

NASA Astrophysics Data System (ADS)

Perez, Camilo; Faust, Belinda; Mehdipour, Ahmad Reza; Francesconi, Kevin A.; Forrest, Lucy R.; Ziegler, Christine

2014-07-01

The Na+-coupled betaine symporter BetP shares a highly conserved fold with other sequence unrelated secondary transporters, for example, with neurotransmitter symporters. Recently, we obtained atomic structures of BetP in distinct conformational states, which elucidated parts of its alternating-access mechanism. Here, we report a structure of BetP in a new outward-open state in complex with an anomalous scattering substrate, adding a fundamental piece to an unprecedented set of structural snapshots for a secondary transporter. In combination with molecular dynamics simulations these structural data highlight important features of the sequential formation of the substrate and sodium-binding sites, in which coordinating water molecules play a crucial role. We observe a strictly interdependent binding of betaine and sodium ions during the coupling process. All three sites undergo progressive reshaping and dehydration during the alternating-access cycle, with the most optimal coordination of all substrates found in the closed state.

Advances in the enzymatic production of L-hexoses.

PubMed

Chen, Ziwei; Zhang, Wenli; Zhang, Tao; Jiang, Bo; Mu, Wanmeng

2016-08-01

Rare sugars have recently drawn attention because of their potential applications and huge market demands in the food and pharmaceutical industries. All L-hexoses are considered rare sugars, as they rarely occur in nature and are thus very expensive. L-Hexoses are important components of biologically relevant compounds as well as being used as precursors for certain pharmaceutical drugs and thus play an important role in the pharmaceutical industry. Many general strategies have been established for the synthesis of L-hexoses; however, the only one used in the biotechnology industry is the Izumoring strategy. In hexose Izumoring, four entrances link the D- to L-enantiomers, ketose 3-epimerases catalyze the C-3 epimerization of L-ketohexoses, and aldose isomerases catalyze the specific bioconversion of L-ketohexoses and the corresponding L-aldohexoses. In this article, recent studies on the enzymatic production of various L-hexoses are reviewed based on the Izumoring strategy.

The two Na+ sites in the human serotonin transporter play distinct roles in the ion coupling and electrogenicity of transport.

PubMed

Felts, Bruce; Pramod, Akula Bala; Sandtner, Walter; Burbach, Nathan; Bulling, Simon; Sitte, Harald H; Henry, L Keith

2014-01-17

Neurotransmitter transporters of the SLC6 family of proteins, including the human serotonin transporter (hSERT), utilize Na(+), Cl(-), and K(+) gradients to induce conformational changes necessary for substrate translocation. Dysregulation of ion movement through monoamine transporters has been shown to impact neuronal firing potentials and could play a role in pathophysiologies, such as depression and anxiety. Despite multiple crystal structures of prokaryotic and eukaryotic SLC transporters indicating the location of both (or one) conserved Na(+)-binding sites (termed Na1 and Na2), much remains uncertain in regard to the movements and contributions of these cation-binding sites in the transport process. In this study, we utilize the unique properties of a mutation of hSERT at a single, highly conserved asparagine on TM1 (Asn-101) to provide several lines of evidence demonstrating mechanistically distinct roles for Na1 and Na2. Mutations at Asn-101 alter the cation dependence of the transporter, allowing Ca(2+) (but not other cations) to functionally replace Na(+) for driving transport and promoting 5-hydroxytryptamine (5-HT)-dependent conformational changes. Furthermore, in two-electrode voltage clamp studies in Xenopus oocytes, both Ca(2+) and Na(+) illicit 5-HT-induced currents in the Asn-101 mutants and reveal that, although Ca(2+) promotes substrate-induced current, it does not appear to be the charge carrier during 5-HT transport. These findings, in addition to functional evaluation of Na1 and Na2 site mutants, reveal separate roles for Na1 and Na2 and provide insight into initiation of the translocation process as well as a mechanism whereby the reported SERT stoichiometry can be obtained despite the presence of two putative Na(+)-binding sites.

The Two Na+ Sites in the Human Serotonin Transporter Play Distinct Roles in the Ion Coupling and Electrogenicity of Transport*

PubMed Central

Felts, Bruce; Pramod, Akula Bala; Sandtner, Walter; Burbach, Nathan; Bulling, Simon; Sitte, Harald H.; Henry, L. Keith

2014-01-01

Neurotransmitter transporters of the SLC6 family of proteins, including the human serotonin transporter (hSERT), utilize Na+, Cl−, and K+ gradients to induce conformational changes necessary for substrate translocation. Dysregulation of ion movement through monoamine transporters has been shown to impact neuronal firing potentials and could play a role in pathophysiologies, such as depression and anxiety. Despite multiple crystal structures of prokaryotic and eukaryotic SLC transporters indicating the location of both (or one) conserved Na+-binding sites (termed Na1 and Na2), much remains uncertain in regard to the movements and contributions of these cation-binding sites in the transport process. In this study, we utilize the unique properties of a mutation of hSERT at a single, highly conserved asparagine on TM1 (Asn-101) to provide several lines of evidence demonstrating mechanistically distinct roles for Na1 and Na2. Mutations at Asn-101 alter the cation dependence of the transporter, allowing Ca2+ (but not other cations) to functionally replace Na+ for driving transport and promoting 5-hydroxytryptamine (5-HT)-dependent conformational changes. Furthermore, in two-electrode voltage clamp studies in Xenopus oocytes, both Ca2+ and Na+ illicit 5-HT-induced currents in the Asn-101 mutants and reveal that, although Ca2+ promotes substrate-induced current, it does not appear to be the charge carrier during 5-HT transport. These findings, in addition to functional evaluation of Na1 and Na2 site mutants, reveal separate roles for Na1 and Na2 and provide insight into initiation of the translocation process as well as a mechanism whereby the reported SERT stoichiometry can be obtained despite the presence of two putative Na+-binding sites. PMID:24293367

Distinguishing Biologically Relevant Hexoses by Water Adduction to the Lithium-Cationized Molecule.

PubMed

Campbell, Matthew T; Chen, Dazhe; Wallbillich, Nicholas J; Glish, Gary L

2017-10-03

A method to distinguish the four most common biologically relevant underivatized hexoses, d-glucose, d-galactose, d-mannose, and d-fructose, using only mass spectrometry with no prior separation/derivatization step has been developed. Electrospray of a solution containing hexose and a lithium salt generates [Hexose+Li] + . The lithium-cationized hexoses adduct water in a quadrupole ion trap. The rate of this water adduction reaction can be used to distinguish the four hexoses. Additionally, for each hexose, multiple lithiation sites are possible, allowing for multiple structures of [Hexose+Li] + . Electrospray produces at least one structure that reacts with water and at least one that does not. The ratio of unreactive lithium-cationized hexose to total lithium-cationized hexose is unique for the four hexoses studied, providing a second method for distinguishing the isomers. Use of the water adduction reaction rate or the unreactive ratio provides two separate methods for confidently (p ≤ 0.02) distinguishing the most common biologically relevant hexoses using only femtomoles of hexose. Additionally, binary mixtures of glucose and fructose were studied. A calibration curve was created by measuring the reaction rate of various samples with different ratios of fructose and glucose. The calibration curve was used to accurately measure the percentage of fructose in three samples of high fructose corn syrup (<4% error).

Bioproduction strategies for rare hexose sugars

NASA Astrophysics Data System (ADS)

Izumori, Ken

2002-03-01

A new strategy for the bioproduction of all ketohexoses was developed using hexitols as intermediates. Biocatalysts used to employ the strategy were D-tagatose 3-epimerase, which epimerizes ketohexoses at the C-3 position, and oxidoreductases, which catalyze oxidation-reduction reactions between ketohexoses and the corresponding hexitols. Arranging all the ketohexoses and hexitols in a symmetric ring and connecting them with 20 biochemical reactions, I was able to construct a design for the bioproduction of all the rare ketohexoses. Various aldose isomerases transform ketohexoses into the corresponding aldohexoses, so the strategy is useful for the bioproduction of all the rare hexose sugars. Furthermore, the design revealed that there are four routes to the L-hexose world from the D-hexose one.

Bioproduction strategies for rare hexose sugars.

PubMed

Izumori, Ken

2002-03-01

A new strategy for the bioproduction of all ketohexoses was developed using hexitols as intermediates. Biocatalysts used to employ the strategy were D-tagatose 3-epimerase, which epimerizes ketohexoses at the C-3 position, and oxidoreductases, which catalyze oxidation-reduction reactions between ketohexoses and the corresponding hexitols. Arranging all the ketohexoses and hexitols in a symmetric ring and connecting them with 20 biochemical reactions, I was able to construct a design for the bioproduction of all the rare ketohexoses. Various aldose isomerases transform ketohexoses into the corresponding aldohexoses, so the strategy is useful for the bioproduction of all the rare hexose sugars. Furthermore, the design revealed that there are four routes to the L-hexose world from the D-hexose one.

Sodium-coupled electrogenic transport of pyroglutamate (5-oxoproline) via SLC5A8, a monocarboxylate transporter.

PubMed

Miyauchi, Seiji; Gopal, Elangovan; Babu, Ellappan; Srinivas, Sonne R; Kubo, Yoshiyuki; Umapathy, Nagavedi S; Thakkar, Santoshanand V; Ganapathy, Vadivel; Prasad, Puttur D

2010-06-01

Pyroglutamate, also known as 5-oxoproline, is a structural analog of proline. This amino acid derivative is a byproduct of glutathione metabolism, and is reabsorbed efficiently in kidney by Na(+)-coupled transport mechanisms. Previous studies have focused on potential participation of amino acid transport systems in renal reabsorption of this compound. Here we show that it is not the amino acid transport systems but instead the Na(+)-coupled monocarboxylate transporter SLC5A8 that plays a predominant role in this reabsorptive process. Expression of cloned human and mouse SLC5A8 in mammalian cells induces Na(+)-dependent transport of pyroglutamate that is inhibitable by various SLC5A8 substrates. SLC5A8-mediated transport of pyroglutamate is saturable with a Michaelis constant of 0.36+/-0.04mM. Na(+)-activation of the transport process exhibits sigmoidal kinetics with a Hill coefficient of 1.8+/-0.4, indicating involvement of more than one Na(+) in the activation process. Expression of SLC5A8 in Xenopuslaevis oocytes induces Na(+)-dependent inward currents in the presence of pyroglutamate under voltage-clamp conditions. The concentration of pyroglutamate necessary for induction of half-maximal current is 0.19+/-0.01mM. The Na(+)-activation kinetics is sigmoidal with a Hill coefficient of 2.3+/-0.2. Ibuprofen, a blocker of SLC5A8, suppressed pyroglutamate-induced currents in SLC5A8-expressing oocytes; the concentration of the blocker necessary for causing half-maximal inhibition is 14+/-1microM. The involvement of SLC5A8 can be demonstrated in rabbit renal brush border membrane vesicles by showing that the Na(+)-dependent uptake of pyroglutamate in these vesicles is inhibitable by known substrates of SLC5A8. The Na(+) gradient-driven pyroglutamate uptake was stimulated by an inside-negative K(+) diffusion potential induced by valinomycin, showing that the uptake process is electrogenic.

Na+/substrate Coupling in the Multidrug Antiporter NorM Probed with a Spin-labeled Substrate

PubMed Central

Steed, P. Ryan; Stein, Richard A.; Mishra, Smriti; Goodman, Michael C.; Mchaourab, Hassane S.

2013-01-01

NorM of the multidrug and toxic compound extrusion (MATE) family of transporters couples the efflux of a broad range of hydrophobic molecules to an inward Na+ gradient across the cell membrane. Several crystal structures of MATE transporters revealed distinct substrate binding sites leading to differing models of the mechanism of ion-coupled substrate extrusion. In the experiments reported here, we observed that a spin-labeled derivative of daunorubicin, Ruboxyl, is transported by NorM from Vibrio cholerae. It is therefore ideal to characterize mechanistically relevant binding interactions with NorM and to directly address the coupling of ion and drug binding. Fluorescence and EPR experiments revealed that Ruboxyl binds to NorM with micromolar affinity and becomes immobilized upon binding, even in the presence of Na+. Using double electron-electron resonance (DEER) spectroscopy, we determined that Ruboxyl binds to a single site on the periplasmic side of the protein. The presence of Na+ did not translocate the substrate to a second site as previously proposed. These experiments surprisingly show that Na+ does not affect the affinity or location of the substrate binding site on detergent-solubilized NorM, thus suggesting that additional factors beyond simple mutual exclusivity of binding, such as the presence of a Na+ gradient across the native membrane, govern Na+/drug coupling during antiport. PMID:23902581

Na+ Interactions with the Neutral Amino Acid Transporter ASCT1*

PubMed Central

Scopelliti, Amanda J.; Heinzelmann, Germano; Kuyucak, Serdar; Ryan, Renae M.; Vandenberg, Robert J.

2014-01-01

The alanine, serine, cysteine transporters (ASCTs) belong to the solute carrier family 1A (SLC1A), which also includes the excitatory amino acid transporters (EAATs) and the prokaryotic aspartate transporter GltPh. Acidic amino acid transport by the EAATs is coupled to the co-transport of three Na+ ions and one proton, and the counter-transport of one K+ ion. In contrast, neutral amino acid exchange by the ASCTs does not require protons or the counter-transport of K+ ions and the number of Na+ ions required is not well established. One property common to SLC1A family members is a substrate-activated anion conductance. We have investigated the number and location of Na+ ions required by ASCT1 by mutating residues in ASCT1 that correspond to residues in the EAATs and GltPh that are involved in Na+ binding. Mutations to all three proposed Na+ sites influence the binding of substrate and/or Na+, or the rate of substrate exchange. A G422S mutation near the Na2 site reduced Na+ affinity, without affecting the rate of exchange. D467T and D467A mutations in the Na1 site reduce Na+ and substrate affinity and also the rate of substrate exchange. T124A and D380A mutations in the Na3 site selectively reduce the affinity for Na+ and the rate of substrate exchange without affecting substrate affinity. In many of the mutants that reduce the rate of substrate transport the amplitudes of the substrate-activated anion conductances are not substantially affected indicating altered ion dependence for channel activation compared with substrate exchange. PMID:24808181

Noncompetitive blocking of human GLUT1 hexose transporter by methylxanthines reveals an exofacial regulatory binding site.

PubMed

Ojeda, Paola; Pérez, Alejandra; Ojeda, Lorena; Vargas-Uribe, Mauricio; Rivas, Coralia I; Salas, Monica; Vera, Juan Carlos; Reyes, Alejandro M

2012-09-01

Glucose transporter (GLUT)1 has become an attractive target to block glucose uptake in malignant cells since most cancer cells overexpress GLUT1 and are sensitive to glucose deprivation. Methylxanthines are natural compounds that inhibit glucose uptake; however, the mechanism of inhibition remains unknown. Here, we used a combination of binding and glucose transport kinetic assays to analyze in detail the effects of caffeine, pentoxifylline, and theophylline on hexose transport in human erythrocytes. The displacement of previously bound cytochalasin B revealed a direct interaction between the methylxanthines and GLUT1. Methylxanthines behave as noncompetitive blockers (inhibition constant values of 2-3 mM) in exchange and zero-trans efflux assays, whereas mixed inhibition with a notable uncompetitive component is observed in zero-trans influx assays (inhibition constant values of 5-12 mM). These results indicate that methylxanthines do not bind to either exofacial or endofacial d-glucose-binding sites but instead interact at a different site accessible by the external face of the transporter. Additionally, infinite-cis exit assays (Sen-Widdas assays) showed that only pentoxifylline disturbed d-glucose for binding to the exofacial substrate site. Interestingly, coinhibition assays showed that methylxanthines bind to a common site on the transporter. We concluded that there is a methylxanthine regulatory site on the external surface of the transporter, which is close but distinguishable from the d-glucose external site. Therefore, the methylxanthine moiety may become an attractive framework for the design of novel specific noncompetitive facilitative GLUT inhibitors.

The Thermoanaerobacter Glycobiome Reveals Mechanisms of Pentose and Hexose Co-Utilization in Bacteria

PubMed Central

Tu, Qichao; Qin, Yujia; Zhou, Aifen; Liu, Wenbin; He, Zhili; Zhou, Jizhong; Xu, Jian

2011-01-01

Thermoanaerobic bacteria are of interest in cellulosic-biofuel production, due to their simultaneous pentose and hexose utilization (co-utilization) and thermophilic nature. In this study, we experimentally reconstructed the structure and dynamics of the first genome-wide carbon utilization network of thermoanaerobes. The network uncovers numerous novel pathways and identifies previously unrecognized but crucial pathway interactions and the associated key junctions. First, glucose, xylose, fructose, and cellobiose catabolism are each featured in distinct functional modules; the transport systems of hexose and pentose are apparently both regulated by transcriptional antiterminators of the BglG family, which is consistent with pentose and hexose co-utilization. Second, glucose and xylose modules cooperate in that the activity of the former promotes the activity of the latter via activating xylose transport and catabolism, while xylose delays cell lysis by sustaining coenzyme and ion metabolism. Third, the vitamin B12 pathway appears to promote ethanologenesis through ethanolamine and 1, 2-propanediol, while the arginine deiminase pathway probably contributes to cell survival in stationary phase. Moreover, by experimentally validating the distinct yet collaborative nature of glucose and xylose catabolism, we demonstrated that these novel network-derived features can be rationally exploited for product-yield enhancement via optimized timing and balanced loading of the carbon supply in a substrate-specific manner. Thus, this thermoanaerobic glycobiome reveals novel genetic features in carbon catabolism that may have immediate industrial implications and provides novel strategies and targets for fermentation and genome engineering. PMID:22022280

Hexose rearrangements upon fragmentation of N-glycopeptides and reductively aminated N-glycans.

PubMed

Wuhrer, Manfred; Koeleman, Carolien A M; Deelder, André M

2009-06-01

Tandem mass spectrometry of glycans and glycoconjugates in protonated form is known to result in rearrangement reactions leading to internal residue loss. Here we studied the occurrence of hexose rearrangements in tandem mass spectrometry of N-glycopeptides and reductively aminated N-glycans by MALDI-TOF/TOF-MS/MS and ESI-ion trap-MS/MS. Fragmentation of proton adducts of oligomannosidic N-glycans of ribonuclease B that were labeled with 2-aminobenzamide and 2-aminobenzoic acid resulted in transfer of one to five hexose residues to the fluorescently tagged innermost N-acetylglucosamine. Glycopeptides from various biological sources with oligomannosidic glycans were likewise shown to undergo hexose rearrangement reactions, resulting in chitobiose cleavage products that have acquired one or two hexose moieties. Tryptic immunoglobulin G Fc-glycopeptides with biantennary N-glycans likewise showed hexose rearrangements resulting in hexose transfer to the peptide moiety retaining the innermost N-acetylglucosamine. Thus, as a general phenomenon, tandem mass spectrometry of reductively aminated glycans as well as glycopeptides may result in hexose rearrangements. This characteristic of glycopeptide MS/MS has to be considered when developing tools for de novo glycopeptide structural analysis.

Involvement of Abscisic Acid in the Coordinated Regulation of a Stress-Inducible Hexose Transporter (VvHT5) and a Cell Wall Invertase in Grapevine in Response to Biotrophic Fungal Infection[W

PubMed Central

Hayes, Matthew A.; Feechan, Angela; Dry, Ian B.

2010-01-01

Biotrophic fungal and oomycete pathogens alter carbohydrate metabolism in infected host tissues. Symptoms such as elevated soluble carbohydrate concentrations and increased invertase activity suggest that a pathogen-induced carbohydrate sink is established. To identify pathogen-induced regulators of carbohydrate sink strength, quantitative real-time polymerase chain reaction was used to measure transcript levels of invertase and hexose transporter genes in biotrophic pathogen-infected grapevine (Vitis vinifera) leaves. The hexose transporter VvHT5 was highly induced in coordination with the cell wall invertase gene VvcwINV by powdery and downy mildew infection. However, similar responses were also observed in response to wounding, suggesting that this is a generalized response to stress. Analysis of the VvHT5 promoter region indicated the presence of multiple abscisic acid (ABA) response elements, suggesting a role for ABA in the transition from source to sink under stress conditions. ABA treatment of grape leaves was found to reproduce the same gene-specific transcriptional changes as observed under biotic and abiotic stress conditions. Furthermore, the key regulatory ABA biosynthetic gene, VvNCED1, was activated under these same stress conditions. VvHT5 promoter::β-glucuronidase-directed expression in transgenic Arabidopsis (Arabidopsis thaliana) was activated by infection with powdery mildew and by ABA treatment, and the expression was closely associated with vascular tissue adjacent to infected regions. Unlike VvHT1 and VvHT3, which appear to be predominantly involved in hexose transport in developing leaves and berries, VvHT5 appears to have a specific role in enhancing sink strength under stress conditions, and this is controlled through ABA. Our data suggest a central role for ABA in the regulation of VvcwINV and VvHT5 expression during the transition from source to sink in response to infection by biotrophic pathogens. PMID:20348211

Human sodium-coupled citrate transporter, the orthologue of Drosophila Indy, as a novel target for lithium action.

PubMed Central

Inoue, Katsuhisa; Zhuang, Lina; Maddox, Dennis M; Smith, Sylvia B; Ganapathy, Vadivel

2003-01-01

NaCT (sodium-coupled citrate transporter) is an Na(+)-coupled citrate transporter identified recently in mammals that mediates the cellular uptake of citrate. It is expressed predominantly in the liver. NaCT is structurally and functionally related to the product of the Indy (I'm not dead yet) gene in Drosophila, the dysfunction of which leads to lifespan extension. Here, we show that NaCT mediates the utilization of extracellular citrate for fat synthesis in human liver cells, and that the process is stimulated by lithium. The transport function of NaCT is enhanced by lithium at concentrations found in humans treated with lithium for bipolar disorders. Valproate and carbamazepine, two other drugs that are used for the treatment of bipolar disorder, do not affect the function of NaCT. The stimulatory effect of Li+ is specific for human NaCT, since NaCTs from other animal species are either inhibited or unaffected by Li+. The data also suggest that two of the four Na(+)-binding sites in human NaCT may become occupied by Li+ to produce the stimulatory effect. The stimulation of NaCT in humans by lithium at therapeutically relevant concentrations has potential clinical implications. We also show here that a single base mutation in codon-500 (TTT-->CTT) in the human NaCT gene, leading to the replacement of phenylalanine with leucine, stimulates the transport function and abolishes the stimulatory effect of lithium. This raises the possibility that genetic mutations in humans may lead to alterations in the constitutive activity of the transporter, with associated clinical consequences. PMID:12826022

Evolved hexose transporter enhances xylose uptake and glucose/xylose co-utilization in Saccharomyces cerevisiae

DOE Office of Scientific and Technical Information (OSTI.GOV)

Reider Apel, Amanda; Ouellet, Mario; Szmidt-Middleton, Heather

Enhancing xylose utilization has been a major focus in Saccharomyces cerevisiae strain-engineering efforts. The incentive for these studies arises from the need to use all sugars in the typical carbon mixtures that comprise standard renewable plant-biomass-based carbon sources. While major advances have been made in developing utilization pathways, the efficient import of five carbon sugars into the cell remains an important bottleneck in this endeavor. Here we use an engineered S. cerevisiae BY4742 strain, containing an established heterologous xylose utilization pathway, and imposed a laboratory evolution regime with xylose as the sole carbon source. We obtained several evolved strains withmore » improved growth phenotypes and evaluated the best candidate using genome resequencing. We observed remarkably few single nucleotide polymorphisms in the evolved strain, among which we confirmed a single amino acid change in the hexose transporter HXT7 coding sequence to be responsible for the evolved phenotype. Lastly, the mutant HXT7(F79S) shows improved xylose uptake rates (Vmax = 186.4 ± 20.1 nmol•min -1•mg -1) that allows the S. cerevisiae strain to show significant growth with xylose as the sole carbon source, as well as partial co-utilization of glucose and xylose in a mixed sugar cultivation.« less

Evolved hexose transporter enhances xylose uptake and glucose/xylose co-utilization in Saccharomyces cerevisiae

DOE PAGES

Reider Apel, Amanda; Ouellet, Mario; Szmidt-Middleton, Heather; ...

2016-01-19

Enhancing xylose utilization has been a major focus in Saccharomyces cerevisiae strain-engineering efforts. The incentive for these studies arises from the need to use all sugars in the typical carbon mixtures that comprise standard renewable plant-biomass-based carbon sources. While major advances have been made in developing utilization pathways, the efficient import of five carbon sugars into the cell remains an important bottleneck in this endeavor. Here we use an engineered S. cerevisiae BY4742 strain, containing an established heterologous xylose utilization pathway, and imposed a laboratory evolution regime with xylose as the sole carbon source. We obtained several evolved strains withmore » improved growth phenotypes and evaluated the best candidate using genome resequencing. We observed remarkably few single nucleotide polymorphisms in the evolved strain, among which we confirmed a single amino acid change in the hexose transporter HXT7 coding sequence to be responsible for the evolved phenotype. Lastly, the mutant HXT7(F79S) shows improved xylose uptake rates (Vmax = 186.4 ± 20.1 nmol•min -1•mg -1) that allows the S. cerevisiae strain to show significant growth with xylose as the sole carbon source, as well as partial co-utilization of glucose and xylose in a mixed sugar cultivation.« less

An Inductive Logic Programming Approach to Validate Hexose Binding Biochemical Knowledge.

PubMed

Nassif, Houssam; Al-Ali, Hassan; Khuri, Sawsan; Keirouz, Walid; Page, David

2010-01-01

Hexoses are simple sugars that play a key role in many cellular pathways, and in the regulation of development and disease mechanisms. Current protein-sugar computational models are based, at least partially, on prior biochemical findings and knowledge. They incorporate different parts of these findings in predictive black-box models. We investigate the empirical support for biochemical findings by comparing Inductive Logic Programming (ILP) induced rules to actual biochemical results. We mine the Protein Data Bank for a representative data set of hexose binding sites, non-hexose binding sites and surface grooves. We build an ILP model of hexose-binding sites and evaluate our results against several baseline machine learning classifiers. Our method achieves an accuracy similar to that of other black-box classifiers while providing insight into the discriminating process. In addition, it confirms wet-lab findings and reveals a previously unreported Trp-Glu amino acids dependency.

Cation Transport Coupled to ATP Hydrolysis by the (Na, K)-ATPase: An Integrated, Animated Model

ERIC Educational Resources Information Center

Leone, Francisco A.; Furriel, Rosa P. M.; McNamara, John C.; Horisberger, Jean D.; Borin, Ivana A.

2010-01-01

An Adobe[R] animation is presented for use in undergraduate Biochemistry courses, illustrating the mechanism of Na[superscript +] and K[superscript +] translocation coupled to ATP hydrolysis by the (Na, K)-ATPase, a P[subscript 2c]-type ATPase, or ATP-powered ion pump that actively translocates cations across plasma membranes. The enzyme is also…

Complete Hexose Isomer Identification with Mass Spectrometry

NASA Astrophysics Data System (ADS)

Nagy, Gabe; Pohl, Nicola L. B.

2015-04-01

The first analytical method is presented for the identification and absolute configuration determination of all 24 aldohexose and 2-ketohexose isomers, including the D and L enantiomers for allose, altrose, galactose, glucose, gulose, idose, mannose, talose, fructose, psicose, sorbose, and tagatose. Two unique fixed ligand kinetic method combinations were discovered to create significant enough energetic differences to achieve chiral discrimination among all 24 hexoses. Each of these 24 hexoses yields unique ratios of a specific pair of fragment ions that allows for simultaneous determination of identification and absolute configuration. This mass spectrometric-based methodology can be readily employed for accurate identification of any isolated monosaccharide from an unknown biological source. This work provides a key step towards the goal of complete de novo carbohydrate analysis.

Active potassium transport coupled to active sodium transport in vesicles reconstituted from purified sodium and potassium ion-activated adenosine triphosphatase from the rectal gland of Squalus acanthias.

PubMed

Hilden, S; Hokin, L E

1975-08-25

Vesicles containing a purified shark rectal gland (sodium + potassium)-activated adenosine triphosphatase-(NaK ATPase) were prepared by dialyzing for 2 days egg lecithin, cholate, and the NaK ATPase purified from the rectal gland of Squalus acanthias. These vesicles were capable of both Na+ and K+ transport. Studies of K+ transport were made by measuring the ATP-stimulated transport outward of 42K+ or 86Rb+. Vesicles were preloaded with isotope by equilibration at 4 degrees for 1 to 3 days. Transport of 42K+ or 86Rb+ was initiated by addition of MgATP to the vesicles. The ATP-dependent exit of either isotope was the same. Experiments are presented which show that this loss of isotope was not due to changes in ion binding but rather due to a loss in the amount of ion trapped in the vesicular volume. The transport of K+ was dependent on external Mg2+. CTP was almost as effective as ATP in stimulating K+ transport, while UTP was relatively ineffective. These effects of nucleotides parallel their effects on Na+ accumulation and their effectiveness as substrates for the enzyme. Potassium transport was inhibited by ouabain and required the presence of Na+. The following asymmetries were seen: (a) addition of external Mg2+ supported K+ transport; (b) ouabain inhibited K+ transport only if it was present inside the vesicles; (c) addition of external Na+ to the vesicles stimulated K+ transport. External Li+ was ineffective as a Na+ substitute. The specific requirement of external Na+ for K+ transport indicates that K+ exit is coupled to Na+ entry. Changes in the internal vesicular ion concentrations were studied with vesicles prepared in 20 mM NaCl and 50 mM KCl. After 1 hour of transport at 25 degrees, a typical Na+ concentration in the vesicles in the presence of ATP was 72 mM. A typical K+ concentration in the vesicles was 10 mM as measured with 42K+ or 6 mM as measured with 86Rb+. The following relationships have been calculated for Na+ transport, K+ transport and ATP

Coupled transport of p-aminohippurate by rat kidney basolateral membrane vesicles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pritchard, J.B.

p-Aminohippuric acid (PAH) transport by basolateral membrane (BLM) vesicles isolated from rat renal cortex was stimulated very little by a Na{sup +} gradient (out > in). However, when micromolar concentrations of glutaric acid or {alpha}-ketoglutaric acid were added in the presence of a out > in Na{sup +} gradient, PAH uptake was accelerated >20-fold and an overshoot of greater than fivefold was produced. Other anions, e.g., fumarate, stimulated PAH uptake very modestly under these conditions, and that stimulation was totally prevented by short circuiting, i.e., with K{sup +} (in = out) and valinomycin. Glutarate-stimulated uptake was inhibited by 4-acetamide-4{prime}-({sup 14}C)-isothiocyanostilbene-2,2{prime}-disulfonicmore » acid (SITS) and probenecid and was slightly stimulated by the imposition of an inside-negative membrane potential. Furthermore, even in the absence of a Na{sup +} gradient, glutarate-loaded vesicles exhibited a marked acceleration of ({sup 3}H)-PAH uptake (5-fold) and a modest overshoot (2.5-fold). These results suggest an indirect coupling of BLM PAH uptake to the Na{sup +} gradient by a cyclic accumulation (Na{sup +}-dependent) of glutarate followed by its efflux from the vesicle in exchange for PAH. This coupled system was absent in apical membranes. Thus net secretory transport of PAH may entail Na{sup +}-dependent, glutarate-driven PAH uptake at the BLM, followed by the exit of PAH into the lumen down its electrochemical gradient, probably in exchange for other anions, e.g., {sup 36}Cl{sup {minus}}, HCO{sub 3}{sup {minus}}, or OH{sup {minus}}.« less

Water transport by Na+-coupled cotransporters of glucose (SGLT1) and of iodide (NIS). The dependence of substrate size studied at high resolution

PubMed Central

Zeuthen*, Thomas; Belhage, Bo; Zeuthen, Emil

2006-01-01

The relation between substrate and water transport was studied in Na+-coupled cotransporters of glucose (SGLT1) and of iodide (NIS) expressed in Xenopus oocytes. The water transport was monitored from changes in oocyte volume at a resolution of 20 pl, more than one order of magnitude better than previous investigations. The rate of cotransport was monitored as the clamp current obtained from two-electrode voltage clamp. The high resolution data demonstrated a fixed ratio between the turn-over of the cotransporter and the rate of water transport. This applied to experiments in which the rate of cotransport was changed by isosmotic application of substrate, by rapid changes in clamp voltage, or by poisoning. Transport of larger substrates gave rise to less water transport. For the rabbit SGLT1, 378 ± 20 (n = 18 oocytes) water molecules were cotransported along with the 2 Na+ ions and the glucose-analogue α-MDG (MW 194); using the larger sugar arbutin (MW 272) this number was reduced by a factor of at least 0.86 ± 0.03 (15). For the human SGLT1 the respective numbers were 234 ± 12 (18) and 0.85 ± 0.8 (7). For NIS, 253 ± 16 (12) water molecules were cotransported for each 2 Na+ and 1 thiocyanate (SCN−, MW 58), with I− as anion (MW 127) only 162 ± 11 (19) water molecules were cotransported. The effect of substrate size suggests a molecular mechanism for water cotransport and is opposite to what would be expected from unstirred layer effects. Data were analysed by a model which combined cotransport and osmosis at the membrane with diffusion in the cytoplasm. The combination of high resolution measurements and precise modelling showed that water transport across the membrane can be explained by cotransport of water in the membrane proteins and that intracellular unstirred layers effects are minute. PMID:16322051

A Conserved Asparagine Residue in Transmembrane Segment 1 (TM1) of Serotonin Transporter Dictates Chloride-coupled Neurotransmitter Transport*

PubMed Central

Henry, L. Keith; Iwamoto, Hideki; Field, Julie R.; Kaufmann, Kristian; Dawson, Eric S.; Jacobs, Miriam T.; Adams, Chelsea; Felts, Bruce; Zdravkovic, Igor; Armstrong, Vanessa; Combs, Steven; Solis, Ernesto; Rudnick, Gary; Noskov, Sergei Y.; DeFelice, Louis J.; Meiler, Jens; Blakely, Randy D.

2011-01-01

Na+- and Cl−-dependent uptake of neurotransmitters via transporters of the SLC6 family, including the human serotonin transporter (SLC6A4), is critical for efficient synaptic transmission. Although residues in the human serotonin transporter involved in direct Cl− coordination of human serotonin transport have been identified, the role of Cl− in the transport mechanism remains unclear. Through a combination of mutagenesis, chemical modification, substrate and charge flux measurements, and molecular modeling studies, we reveal an unexpected role for the highly conserved transmembrane segment 1 residue Asn-101 in coupling Cl− binding to concentrative neurotransmitter uptake. PMID:21730057

Insights into the ion-coupling mechanism in the MATE transporter NorM-VC

NASA Astrophysics Data System (ADS)

Krah, Alexander; Zachariae, Ulrich

2017-08-01

Bacteria have developed a variety of different mechanisms to defend themselves from compounds that are toxic to them, such as antibiotics. One of these defence mechanisms is the expulsion of drugs or other noxious compounds by multidrug efflux pumps. Multidrug and toxic compound extrusion (MATE) transporters are efflux pumps that extrude metabolic waste and a variety of antibiotics out of the cell, using an ion gradient as energy source. They function via an alternating-access mechanism. When ions bind in the outward facing conformation, a large conformational change to the inward facing conformation is induced, from which the ion is released and the extruded chemical compound is bound. NorM proteins, which are usually coupled to a Na+ gradient, are members of the MATE family. However, for NorM-VC from Vibrio cholerae, it has been shown that this MATE transporter is additionally coupled to protons. How H+ and Na+ binding are coupled mechanistically to enable drug antiport is not well understood. In this study, we use molecular dynamics simulations to illuminate the sequence of ion binding events that enable efflux. Understanding this antiport mechanism is important to support the development of novel compounds that specifically inhibit the functional cycle of NorM transporters.

Nutritional Relationships in Schistosomiasis.

DTIC Science & Technology

1978-07-01

to presence of ou~bain, phlorizin, phloretin . 1lexose mediated accumulation is Na coupled. Accession For NTIS GREA&I DDC TAB Unannounced...mediat,!d system is apparently Na + dependent and sensitive to presence of ouabain, phlorizin, phloretin . Hexose mediated accumulation is Na+ coupled. i...decreased Na+, ouabain, phiorizin phloretin and other sugars. Studies with Na22 suggest that the accumulation of 2-deoxyglucose and Na+ were coupled

Insulin stimulation of glucose transport in isolated rat adipocytes. Functional evidence for insulin activation of intrinsic transporter activity within the plasma membrane.

PubMed Central

Hyslop, P A; Kuhn, C E; Sauerheber, R D

1985-01-01

We examined the effects of the membrane-impermeant amino-group-modifying agent fluorescein isothiocyanate (FITC) on the basal and insulin-stimulated hexose-transport activity of isolated rat adipocytes. Pre-treatment of cells with FITC causes irreversible inhibition of transport measured in subsequently washed cells. Transport activity was inhibited by approx. 50% with 2 mM-FITC in 8 min. The cells respond to insulin, after FITC treatment and removal, and the fold increase in transport above the basal value caused by maximal concentrations of insulin was independent of the concentration of FITC used for pre-treatment over the range 0-2 mM, where basal activity was progressively inhibited. The ability of FITC to modify selectively hexose transporters accessible only to the external milieu was evaluated by two methods. (1) Free intracellular FITC, and the distribution of FITC bound to cellular components, were assessed after dialysis of the homogenate and subcellular fractionation on sucrose gradients by direct spectroscopic measurement of fluorescein. Most (98%) of the FITC was associated with the non-diffusible fractions. Equilibrium sucrose-density-gradient centrifugation of the homogenate demonstrated that the subcellular distribution of the bound FITC correlated with the density distribution of a plasma-membrane marker, but not markers for Golgi, endoplasmic reticulum, mitochondria or protein. Exposing the cellular homogenate, rather than the intact cell preparation, to 2 mM-FITC resulted in a 4-5-fold increase in total bound FITC, and the density-distribution profile more closely resembled the distribution of total protein. (2) Incubation of hexokinase preparations with FITC rapidly and irreversibly inactivates this protein. However, both intracellular hexokinase total activity and its apparent Michaelis constant for glucose were unaffected in FITC-treated intact cells. Further control experiments demonstrated that FITC pre-treatment of cells had no effect on

CryoEM structure of the human SLC4A4 sodium-coupled acid-base transporter NBCe1.

PubMed

Huynh, Kevin W; Jiang, Jiansen; Abuladze, Natalia; Tsirulnikov, Kirill; Kao, Liyo; Shao, Xuesi; Newman, Debra; Azimov, Rustam; Pushkin, Alexander; Zhou, Z Hong; Kurtz, Ira

2018-03-02

Na + -coupled acid-base transporters play essential roles in human biology. Their dysfunction has been linked to cancer, heart, and brain disease. High-resolution structures of mammalian Na + -coupled acid-base transporters are not available. The sodium-bicarbonate cotransporter NBCe1 functions in multiple organs and its mutations cause blindness, abnormal growth and blood chemistry, migraines, and impaired cognitive function. Here, we have determined the structure of the membrane domain dimer of human NBCe1 at 3.9 Å resolution by cryo electron microscopy. Our atomic model and functional mutagenesis revealed the ion accessibility pathway and the ion coordination site, the latter containing residues involved in human disease-causing mutations. We identified a small number of residues within the ion coordination site whose modification transformed NBCe1 into an anion exchanger. Our data suggest that symporters and exchangers utilize comparable transport machinery and that subtle differences in their substrate-binding regions have very significant effects on their transport mode.

Quantifying the Contribution of Grape Hexoses to Wine Volatiles by High-Precision [U13C]-Glucose Tracer Studies

PubMed Central

Nisbet, Mark A.; Tobias, Herbert J.; Brenna, J. Thomas; Sacks, Gavin L.; Mansfield, Anna Katharine

2016-01-01

Many fermentation volatiles important to wine aroma potentially arise from yeast metabolism of hexose sugars, but assessing the relative importance of these pathways is challenging due to high endogenous hexose substrate concentrations. To overcome this problem, gas chromatography combustion isotope ratio mass spectrometry (GC-C-IRMS) was used to measure high-precision 13C/12C isotope ratios of volatiles in wines produced from juices spiked with tracer levels (0.01–1 APE) of uniformly labeled [U-13C]-glucose. The contribution of hexose to individual volatiles was determined from the degree of 13C enrichment. As expected, straight-chain fatty acids and their corresponding ethyl esters were derived almost exclusively from hexoses. Most fusel alcohols and their acetate esters were also majority hexose-derived, indicating the importance of anabolic pathways for their formation. Only two compounds were not derived primarily from hexoses (hexanol and isobutyric acid). This approach can be extended to other food systems or substrates for studying precursor–product relationships. PMID:24960193

Quantifying the contribution of grape hexoses to wine volatiles by high-precision [U¹³C]-glucose tracer studies.

PubMed

Nisbet, Mark A; Tobias, Herbert J; Brenna, J Thomas; Sacks, Gavin L; Mansfield, Anna Katharine

2014-07-16

Many fermentation volatiles important to wine aroma potentially arise from yeast metabolism of hexose sugars, but assessing the relative importance of these pathways is challenging due to high endogenous hexose substrate concentrations. To overcome this problem, gas chromatography combustion isotope ratio mass spectrometry (GC-C-IRMS) was used to measure high-precision (13)C/(12)C isotope ratios of volatiles in wines produced from juices spiked with tracer levels (0.01-1 APE) of uniformly labeled [U-(13)C]-glucose. The contribution of hexose to individual volatiles was determined from the degree of (13)C enrichment. As expected, straight-chain fatty acids and their corresponding ethyl esters were derived almost exclusively from hexoses. Most fusel alcohols and their acetate esters were also majority hexose-derived, indicating the importance of anabolic pathways for their formation. Only two compounds were not derived primarily from hexoses (hexanol and isobutyric acid). This approach can be extended to other food systems or substrates for studying precursor-product relationships.

IMMUNOCYTOCHEMICAL IDENTIFICATION OF ELECTRONEUTRAL NA+-COUPLED HCO3− TRANSPORTERS IN FRESHLY DISSOCIATED MOUSE MEDULLARY RAPHÉ NEURONS

PubMed Central

COLEY, A. A.; RUFFIN, V. A.; MOSS, F. J.; HOPFER, U.; BORON, W. F.

2013-01-01

The medullary raphé (MR) of the medulla oblongata contains chemosensitive neurons that respond to increases in arterial [CO2], by altering firing rate, with increases being associated with serotonergic (5-hydroxytryptamine [5HT]) neurons and decreases, with GABAergic neurons. Both types of neurons contribute to increased alveolar ventilation. Decreases in intracellular pH are thought to link the rise in [CO2] to increased ventilation. Because electroneutral Na+-coupled HCO3− transporters (nNCBTs), which help protect cells from intracellular acidosis, are expressed robustly in the neurons of the central nervous system, a key question is whether these transporters are present in chemosensitive neurons. Therefore, we used an immunocytochemistry approach to identify neurons (using a microtubule associated protein-2 monoclonal antibody) and specifically 5HT neurons (TPH monoclonal antibody) or GABAergic neurons (GAD2 monoclonal antibody) in freshly dissociated cells from the mouse MR. We also co-labeled with polyclonal antibodies against the three nNCBTs: NBCn1, NDCBE, and NBCn2. We exploited ePet-EYFP (enhanced yellow fluorescent protein) mice (with EYFP-labeled 5HT neurons) as well as mice genetically deficient in each of the three nNCBTs. Quantitative image analysis distinguished positively stained cells from background signals. We found that >80% of GAD2+ cells also were positive for NDCBE, and >90% of the TPH+ and GAD2+ cells were positive for the other nNCBTs. Assuming that the transporters are independently distributed among neurons, we can conclude that virtually all chemosensitive MR neurons contain at least one nNCBT. PMID:23500099

Na-coupled bicarbonate transporters of the Slc4 family in the nervous system: function, localization, and relevance to neurologic function

PubMed Central

Majumdar, Debeshi; Bevensee, Mark O.

2010-01-01

Many cellular processes including neuronal activity are sensitive to changes in intracellular and/or extracellular pH— both of which are regulated by acid-base transporter activity. HCO3−-dependent transporters are particularly potent regulators of intracellular pH in neurons and astrocytes, and also contribute to the composition of the cerebrospinal fluid (CSF). The molecular physiology of HCO3− transporters has advanced considerably over the past ~14 years as investigators have cloned and characterized the function and localization of many Na-Coupled Bicarbonate Transporters of the Slc4 family (NCBTs). In this review, we provide an updated overview of the function and localization of NCBTs in the nervous system. Multiple NCBTs are expressed in neurons and astrocytes in various brain regions, as well as in epithelial cells of the choroid plexus. Characteristics of human patients with SLC4 gene mutations/deletions and results from recent studies on mice with Slc4 gene disruptions highlight the functional importance of NCBTs in neuronal activity, somatosensory function, and CSF production. Furthermore, energy-deficient states (e.g., hypoxia and ischemia) lead to altered expression and activity of NCBTs. Thus, recent studies expand our understanding of the role of NCBTs in regulating the pH and ionic composition of the nervous system that can modulate neuronal activity. PMID:20884330

Interaction of α-Lipoic Acid with the Human Na+/Multivitamin Transporter (hSMVT)*

PubMed Central

Zehnpfennig, Britta; Wiriyasermkul, Pattama; Carlson, David A.; Quick, Matthias

2015-01-01

The human Na+/multivitamin transporter (hSMVT) has been suggested to transport α-lipoic acid (LA), a potent antioxidant and anti-inflammatory agent used in therapeutic applications, e.g. in the treatment of diabetic neuropathy and Alzheimer disease. However, the molecular basis of the cellular delivery of LA and in particular the stereospecificity of the transport process are not well understood. Here, we expressed recombinant hSMVT in Pichia pastoris and used affinity chromatography to purify the detergent-solubilized protein followed by reconstitution of hSMVT in lipid bilayers. Using a combined approach encompassing radiolabeled LA transport and equilibrium binding studies in conjunction with the stabilized R-(+)- and S-(−)-enantiomers and the R,S-(+/−) racemic mixture of LA or lipoamide, we identified the biologically active form of LA, R-LA, to be the physiological substrate of hSMVT. Interaction of R-LA with hSMVT is strictly dependent on Na+. Under equilibrium conditions, hSMVT can simultaneously bind ∼2 molecules of R-LA in a biphasic binding isotherm with dissociation constants (Kd) of 0.9 and 7.4 μm. Transport of R-LA in the oocyte and reconstituted system is exclusively dependent on Na+ and exhibits an affinity of ∼3 μm. Measuring transport with known amounts of protein in proteoliposomes containing hSMVT in outside-out orientation yielded a catalytic turnover number (kcat) of about 1 s−1, a value that is well in agreement with other Na+-coupled transporters. Our data suggest that hSMVT-mediated transport is highly specific for R-LA at our tested concentration range, a finding with wide ramifications for the use of LA in therapeutic applications. PMID:25971966

NMR studies on Na+ transport in Synechococcus PCC 6311

NASA Technical Reports Server (NTRS)

Nitschmann, W. H.; Packer, L.

1992-01-01

The freshwater cyanobacterium Synechococcus PCC 6311 is able to adapt to grow after sudden exposure to salt (NaCl) stress. We have investigated the mechanism of Na+ transport in these cells during adaptation to high salinity. Na+ influx under dark aerobic conditions occurred independently of delta pH or delta psi across the cytoplasmic membrane, ATPase activity, and respiratory electron transport. These findings are consistent with the existence of Na+/monovalent anion cotransport or simultaneous Na+/H+ +anion/OH- exchange. Na+ influx was dependent on Cl-, Br-, NO3-, or NO2-. No Na+ uptake occurred after addition of NaI, NaHCO3, or Na2SO4. Na+ extrusion was absolutely dependent on delta pH and on an ATPase activity and/or on respiratory electron transport. This indicates that Na+ extrusion via Na+/H+ exchange is driven by primary H+ pumps in the cytoplasmic membrane. Cells grown for 4 days in 0.5 m NaCl medium, "salt-grown cells," differ from control cells by a lower maximum velocity of Na+ influx and by lower steady-state ratios of [Na+]in/[Na+]out. These results indicate that cells grown in high-salt medium increase their capacity to extrude Na+. During salt adaptation Na+ extrusion driven by respiratory electron transport increased from about 15 to 50%.

Effects of topical application of aqueous solutions of hexoses on epidermal permeability barrier recovery rate after barrier disruption.

PubMed

Denda, Mitsuhiro

2011-11-01

Previous studies have suggested that hexose molecules influence the stability of phospholipid bilayers. Therefore, the effects of topical application of all 12 stereoisomers of dextro-hexose on the epidermal barrier recovery rate after barrier disruption were evaluated. Immediately after tape stripping, 0.1 m aqueous solution of each hexose was applied on hairless mouse skin. Among the eight dextro-aldohexoses, topical application of altose, idose, mannose and talose accelerated the barrier recovery, while allose, galactose, glucose and gulose had no effect. Among the four dextro-ketohexoses, psicose, fructose, sorbose and tagatose all accelerated the barrier recovery. As the effects of hexoses on the barrier recovery rate appeared within 1 h, the mechanism is unlikely to be genomic. Instead, these hexoses may influence phase transition of the lipid bilayers of lamellar bodies and cell membrane, a crucial step in epidermal permeability barrier homeostasis. © 2011 John Wiley & Sons A/S.

Multidrug Transport Protein NorM from Vibrio cholerae Simultaneously Couples to Sodium- and Proton-Motive Force*

PubMed Central

Jin, Yoonhee; Nair, Asha; van Veen, Hendrik W.

2014-01-01

Membrane transporters belonging to the multidrug and toxic compound extrusion family mediate the efflux of unrelated pharmaceuticals from the interior of the cell in organisms ranging from bacteria to human. These proteins are thought to fall into two classes that couple substrate efflux to the influx of either Na+ or H+. We studied the energetics of drug extrusion by NorM from Vibrio cholerae in proteoliposomes in which purified NorM protein was functionally reconstituted in an inside-out orientation. We establish that NorM simultaneously couples to the sodium-motive force and proton-motive force, and biochemically identify protein regions and residues that play important roles in Na+ or H+ binding. As the positions of protons are not available in current medium and high-resolution crystal structures of multidrug and toxic compound extrusion transporters, our findings add a previously unrecognized parameter to mechanistic models based of these structures. PMID:24711447

The position of an arginine residue influences substrate affinity and K+ coupling in the human glutamate transporter, EAAT1.

PubMed

Ryan, Renae M; Kortt, Nicholas C; Sirivanta, Tan; Vandenberg, Robert J

2010-07-01

Glutamate is the predominant excitatory neurotransmitter in the mammalian central nervous system and extracellular glutamate levels are controlled by a family of transporters known as excitatory amino acid transporters (EAATs). The EAATs transport glutamate and aspartate with similar micromolar affinities and this transport is coupled to the movement of Na(+), K(+), and H(+). The crystal structure of a prokaryotic homologue of the EAATs, aspartate transporter from Pyrococcus horokoshii (Glt(Ph)), has yielded important insights into the architecture of this transporter family. Glt(Ph) is a Na(+)-dependent transporter that has significantly higher affinity for aspartate over glutamate and is not coupled to H(+) or K(+). The highly conserved carboxy-terminal domains of the EAATs and Glt(Ph) contain the substrate and ion binding sites, however, there are a couple of striking differences in this region that we have investigated to better understand the transport mechanism. An arginine residue is in close proximity to the substrate binding site of both Glt(Ph) and the EAATs, but is located in transmembrane domain (TM) 8 in the EAATs and hairpin loop 1 (HP1) of Glt(Ph). Here we report that the position of this arginine residue can explain some of the functional differences observed between the EAATs and Glt(Ph). Moving the arginine residue from TM8 to HP1 in EAAT1 results in a transporter that has significantly increased affinity for both glutamate and aspartate and is K(+) independent. Conversely, moving the arginine residue from HP1 to TM8 in Glt(Ph) results in a transporter that has reduced affinity for aspartate.

Plasmodial sugar transporters as anti-malarial drug targets and comparisons with other protozoa.

PubMed

Slavic, Ksenija; Krishna, Sanjeev; Derbyshire, Elvira T; Staines, Henry M

2011-06-15

Glucose is the primary source of energy and a key substrate for most cells. Inhibition of cellular glucose uptake (the first step in its utilization) has, therefore, received attention as a potential therapeutic strategy to treat various unrelated diseases including malaria and cancers. For malaria, blood forms of parasites rely almost entirely on glycolysis for energy production and, without energy stores, they are dependent on the constant uptake of glucose. Plasmodium falciparum is the most dangerous human malarial parasite and its hexose transporter has been identified as being the major glucose transporter. In this review, recent progress regarding the validation and development of the P. falciparum hexose transporter as a drug target is described, highlighting the importance of robust target validation through both chemical and genetic methods. Therapeutic targeting potential of hexose transporters of other protozoan pathogens is also reviewed and discussed.

Regulation of Epithelial Sodium Transport via Epithelial Na+ Channel

PubMed Central

Marunaka, Yoshinori; Niisato, Naomi; Taruno, Akiyuki; Ohta, Mariko; Miyazaki, Hiroaki; Hosogi, Shigekuni; Nakajima, Ken-ichi; Kusuzaki, Katsuyuki; Ashihara, Eishi; Nishio, Kyosuke; Iwasaki, Yoshinobu; Nakahari, Takashi; Kubota, Takahiro

2011-01-01

Renal epithelial Na+ transport plays an important role in homeostasis of our body fluid content and blood pressure. Further, the Na+ transport in alveolar epithelial cells essentially controls the amount of alveolar fluid that should be kept at an appropriate level for normal gas exchange. The epithelial Na+ transport is generally mediated through two steps: (1) the entry step of Na+ via epithelial Na+ channel (ENaC) at the apical membrane and (2) the extrusion step of Na+ via the Na+, K+-ATPase at the basolateral membrane. In general, the Na+ entry via ENaC is the rate-limiting step. Therefore, the regulation of ENaC plays an essential role in control of blood pressure and normal gas exchange. In this paper, we discuss two major factors in ENaC regulation: (1) activity of individual ENaC and (2) number of ENaC located at the apical membrane. PMID:22028593

Quantification of total hexose on dry blood spot by tandem mass spectrometry.

PubMed

Gong, Zhenhua; Tian, Guoli; Huang, Qiwei; Wang, Yanmin; Ge, Qingwei

2012-12-01

Because hypoglycemia and hyperglycemia are harmful and not always associated with overt clinical signs, it is necessary to have methods available to screen for glucose levels to detect hypoglycemia and diabetes as early as possible. A new method for such screening and the clinical determination of blood total hexose on a dry blood spot (DBS) using tandem mass spectrometry (MS/MS) was developed. The serum glucose controls and blood were prepared as DBS and then extracted into a methanol solution containing isotope-labeled internal standards. The methanolic extraction was subjected to HPLC, followed by MS/MS in positive ion mode. Multiple-reaction monitoring of m/z 203.1→23 was used to detect hexose, and m/z 209.0→23 was used for 13C6-D-glucose. The recoveries of blood glucose by MS/MS were 90%-102% with an R(2) value of 0.999 after linear regression (p<0.001). The controls were within an acceptable range, and the coefficients of variation were less than 10%. The blood total hexose in neonates aged 3-7 days (6.41±1.46 mmol/L) was lower than that in neonates aged 8-30 days (6.66±1.38 mmol/L), and it was lower in neonates than in children aged 1-72 months (7.19±1.87 mmol/L). Quantification of total hexose on a dry blood spot by MS/MS is accurate, reliable and feasible for screening and clinical tests. Copyright © 2012 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Multidrug transport protein norM from vibrio cholerae simultaneously couples to sodium- and proton-motive force.

PubMed

Jin, Yoonhee; Nair, Asha; van Veen, Hendrik W

2014-05-23

Membrane transporters belonging to the multidrug and toxic compound extrusion family mediate the efflux of unrelated pharmaceuticals from the interior of the cell in organisms ranging from bacteria to human. These proteins are thought to fall into two classes that couple substrate efflux to the influx of either Na(+) or H(+). We studied the energetics of drug extrusion by NorM from Vibrio cholerae in proteoliposomes in which purified NorM protein was functionally reconstituted in an inside-out orientation. We establish that NorM simultaneously couples to the sodium-motive force and proton-motive force, and biochemically identify protein regions and residues that play important roles in Na(+) or H(+) binding. As the positions of protons are not available in current medium and high-resolution crystal structures of multidrug and toxic compound extrusion transporters, our findings add a previously unrecognized parameter to mechanistic models based of these structures. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

The intracellular Na(+)/H(+) exchanger NHE7 effects a Na(+)-coupled, but not K(+)-coupled proton-loading mechanism in endocytosis.

PubMed

Milosavljevic, Nina; Monet, Michaël; Léna, Isabelle; Brau, Frédéric; Lacas-Gervais, Sandra; Feliciangeli, Sylvain; Counillon, Laurent; Poët, Mallorie

2014-05-08

Vesicular H(+)-ATPases and ClC-chloride transporters are described to acidify intracellular compartments, which also express the highly conserved Na(+)/H(+) exchangers NHE6, NHE7, and NHE9. Mutations of these exchangers cause autism-spectrum disorders and neurodegeneration. NHE6, NHE7, and NHE9 are hypothesized to exchange cytosolic K(+) for H(+) and alkalinize vesicles, but this notion has remained untested in K(+) because their intracellular localization prevents functional measurements. Using proton-killing techniques, we selected a cell line that expresses wild-type NHE7 at the plasma membrane, enabling measurement of the exchanger's transport parameters. We found that NHE7 transports Li(+) and Na(+), but not K(+), is nonreversible in physiological conditions and is constitutively activated by cytosolic H(+). Therefore, NHE7 acts as a proton-loading transporter rather than a proton leak. NHE7 mediates an acidification of intracellular vesicles that is additive to that of V-ATPases and that accelerates endocytosis. This study reveals an unexpected function for vesicular Na(+)/H(+) exchangers and provides clues for understanding NHE-linked neurological disorders. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Sodium-coupled neutral amino acid (System N/A) transporters of the SLC38 gene family.

PubMed

Mackenzie, Bryan; Erickson, Jeffrey D

2004-02-01

The sodium-coupled neutral amino acid transporters (SNAT) of the SLC38 gene family resemble the classically-described System A and System N transport activities in terms of their functional properties and patterns of regulation. Transport of small, aliphatic amino acids by System A subtypes (SNAT1, SNAT2, and SNAT4) is rheogenic and pH sensitive. The System N subtypes SNAT3 and SNAT5 also countertransport H(+), which may be key to their operation in reverse, and have narrower substrate profiles than do the System A subtypes. Glutamine emerges as a favored substrate throughout the family, except for SNAT4. The SLC38 transporters undoubtedly play many physiological roles including the transfer of glutamine from astrocyte to neuron in the CNS, ammonia detoxification and gluconeogenesis in the liver, and the renal response to acidosis. Probing their regulation has revealed additional roles, and recent work has considered SLC38 transporters as therapeutic targets in neoplasia.

Plasmodial sugar transporters as anti-malarial drug targets and comparisons with other protozoa

PubMed Central

2011-01-01

Glucose is the primary source of energy and a key substrate for most cells. Inhibition of cellular glucose uptake (the first step in its utilization) has, therefore, received attention as a potential therapeutic strategy to treat various unrelated diseases including malaria and cancers. For malaria, blood forms of parasites rely almost entirely on glycolysis for energy production and, without energy stores, they are dependent on the constant uptake of glucose. Plasmodium falciparum is the most dangerous human malarial parasite and its hexose transporter has been identified as being the major glucose transporter. In this review, recent progress regarding the validation and development of the P. falciparum hexose transporter as a drug target is described, highlighting the importance of robust target validation through both chemical and genetic methods. Therapeutic targeting potential of hexose transporters of other protozoan pathogens is also reviewed and discussed. PMID:21676209

Role of Secondary Transporters and Phosphotransferase Systems in Glucose Transport by Oenococcus oeni ▿

PubMed Central

Kim, Ok Bin; Richter, Hanno; Zaunmüller, Tanja; Graf, Sabrina; Unden, Gottfried

2011-01-01

Glucose uptake by the heterofermentative lactic acid bacterium Oenococcus oeni B1 was studied at the physiological and gene expression levels. Glucose- or fructose-grown bacteria catalyzed uptake of [14C]glucose over a pH range from pH 4 to 9, with maxima at pHs 5.5 and 7. Uptake occurred in two-step kinetics in a high- and low-affinity reaction. The high-affinity uptake followed Michaelis-Menten kinetics and required energization. It accumulated the radioactivity of glucose by a factor of 55 within the bacteria. A large portion (about 80%) of the uptake of glucose was inhibited by protonophores and ionophores. Uptake of the glucose at neutral pH was not sensitive to degradation of the proton potential, Δp. Expression of the genes OEOE_0819 and OEOE_1574 (here referred to as 0819 and 1574), coding for secondary transporters, was induced by glucose as identified by quantitative real-time (RT)-PCR. The genes 1574 and 0819 were able to complement growth of a Bacillus subtilis hexose transport-deficient mutant on glucose but not on fructose. The genes 1574 and 0819 therefore encode secondary transporters for glucose, and the transports are presumably Δp dependent. O. oeni codes, in addition, for a phosphotransferase transport system (PTS) (gene OEOE_0464 [0464] for the permease) with similarity to the fructose- and mannose-specific PTS of lactic acid bacteria. Quantitative RT-PCR showed induction of the gene 0464 by glucose and by fructose. The data suggest that the PTS is responsible for Δp-independent hexose transport at neutral pH and for the residual Δp-independent transport of hexoses at acidic pH. PMID:22020640

Transepithelial glucose transport and Na+/K+ homeostasis in enterocytes: an integrative model

PubMed Central

Drengstig, Tormod; Ruoff, Peter

2014-01-01

The uptake of glucose and the nutrient coupled transcellular sodium traffic across epithelial cells in the small intestine has been an ongoing topic in physiological research for over half a century. Driving the uptake of nutrients like glucose, enterocytes must have regulatory mechanisms that respond to the considerable changes in the inflow of sodium during absorption. The Na-K-ATPase membrane protein plays a major role in this regulation. We propose the hypothesis that the amount of active Na-K-ATPase in enterocytes is directly regulated by the concentration of intracellular Na+ and that this regulation together with a regulation of basolateral K permeability by intracellular ATP gives the enterocyte the ability to maintain ionic Na+/K+ homeostasis. To explore these regulatory mechanisms, we present a mathematical model of the sodium coupled uptake of glucose in epithelial enterocytes. Our model integrates knowledge about individual transporter proteins including apical SGLT1, basolateral Na-K-ATPase, and GLUT2, together with diffusion and membrane potentials. The intracellular concentrations of glucose, sodium, potassium, and chloride are modeled by nonlinear differential equations, and molecular flows are calculated based on experimental kinetic data from the literature, including substrate saturation, product inhibition, and modulation by membrane potential. Simulation results of the model without the addition of regulatory mechanisms fit well with published short-term observations, including cell depolarization and increased concentration of intracellular glucose and sodium during increased concentration of luminal glucose/sodium. Adding regulatory mechanisms for regulation of Na-K-ATPase and K permeability to the model show that our hypothesis predicts observed long-term ionic homeostasis. PMID:24898586

Na+-independent phosphate transport in Caco2BBE cells

PubMed Central

Candeal, Eduardo; Caldas, Yupanqui A.; Guillén, Natalia; Levi, Moshe

2014-01-01

Pi transport in epithelia has both Na+-dependent and Na+-independent components, but so far only Na+-dependent transporters have been characterized in detail and molecularly identified. Consequently, in the present study, we initiated the characterization and analysis of intestinal Na+-independent Pi transport using an in vitro model, Caco2BBE cells. Only Na+-independent Pi uptake was observed in these cells, and Pi uptake was dramatically increased when cells were incubated in high-Pi DMEM (4 mM) from 1 day to several days. No response to low-Pi medium was observed. The increased Pi transport was mainly caused by Vmax changes, and it was prevented by actinomycin D and cycloheximide. Pi transport in cells grown in 1 mM Pi (basal DMEM) decreased at pH > 7.5, and it was inhibited with proton ionophores. Pi transport in cells incubated with 4 mM Pi increased with alkaline pH, suggesting a preference for divalent phosphate. Pi uptake in cells in 1 mM Pi was completely inhibited only by Pi and partially inhibited by phosphonoformate, oxalate, DIDS, SITS, SO42−, HCO3−, and arsenate. This inhibition pattern suggests that more than one Pi transporter is active in cells maintained with 1 mM Pi. Phosphate transport from cells maintained at 4 mM Pi was only partially inhibited by phosphonoformate, oxalate, and arsenate. Attempts to identify the responsible transporters showed that multifunctional anion exchangers of the Slc26 family as well as members of Slc17, Slc20, and Slc37 and the Pi exporter xenotropic and polytropic retrovirus receptor 1 are not involved. PMID:25298422

Na+-dependent and Na+-independent betaine transport across the apical membrane of rat renal epithelium.

PubMed

Cano, Mercedes; Calonge, María L; Ilundáin, Anunciación A

2015-10-01

The low renal excretion of betaine indicates that the kidney efficiently reabsorbs the betaine filtered by the glomeruli but the mechanisms involved in such a process have been scarcely investigated. We have detected concentrative and non-concentrative betaine transport activity in brush-border membrane vesicles (BBMV) from rat renal cortex and medulla. The concentrative system is the Sodium/Imino-acid Transporter 1 (SIT1) because it is Na+- and Cl--dependent, electrogenic and is inhibited by an anti-SIT1 antibody. Its apparent affinity constant for betaine, Kt, is 1.1±0.5 mM and its maximal transport velocity, Vmax, 0.5±0.1 nmol betaine/mg protein/s. Inhibitors of the Na+/Cl-/betaine uptake are L-proline (75%) and cold betaine, L-carnitine and choline (40-60%). Neither creatine, TEA, taurine, β-alanine, GABA nor glycine significantly inhibited Na+/Cl-/betaine uptake. The non-concentrative betaine transport system is Na+- and H+-independent, electroneutral, with a Kt for betaine of 47±7 μM and a Vmax of 7.8±1 pmol betaine/mg protein/s. Its transport activity is nearly abolished by betaine, followed by L-carnitine (70-80%) and proline (40-50%), but a difference from the Na+/Cl-/betaine transport is that it is inhibited by TEA (approx. 50%) and unaffected by choline. The underlying carrier functions as an antiporter linking betaine entry into the BBMV with the efflux of either L-carnitine or betaine, an exchange unaffected by the anti-SIT1 antibody. As far as we know this is the first work reporting that betaine crosses the apical membrane of rat renal epithelium by SIT1 and by a Na+- and H+-independent transport system. Copyright © 2015 Elsevier B.V. All rights reserved.

Ameloblast Modulation and Transport of Cl−, Na+, and K+ during Amelogenesis

PubMed Central

Bronckers, A.L.J.J.; Lyaruu, D.; Jalali, R.; Medina, J.F.; Zandieh-Doulabi, B.; DenBesten, P.K.

2015-01-01

Ameloblasts express transmembrane proteins for transport of mineral ions and regulation of pH in the enamel space. Two major transporters recently identified in ameloblasts are the Na+K+-dependent calcium transporter NCKX4 and the Na+-dependent HPO42– (Pi) cotransporter NaPi-2b. To regulate pH, ameloblasts express anion exchanger 2 (Ae2a,b), chloride channel Cftr, and amelogenins that can bind protons. Exposure to fluoride or null mutation of Cftr, Ae2a,b, or Amelx each results in formation of hypomineralized enamel. We hypothesized that enamel hypomineralization associated with disturbed pH regulation results from reduced ion transport by NCKX4 and NaPi-2b. This was tested by correlation analyses among the levels of Ca, Pi, Cl, Na, and K in forming enamel of mice with null mutation of Cftr, Ae2a,b, and Amelx, according to quantitative x-ray electron probe microanalysis. Immunohistochemistry, polymerase chain reaction analysis, and Western blotting confirmed the presence of apical NaPi-2b and Nckx4 in maturation-stage ameloblasts. In wild-type mice, K levels in enamel were negatively correlated with Ca and Cl but less negatively or even positively in fluorotic enamel. Na did not correlate with P or Ca in enamel of wild-type mice but showed strong positive correlation in fluorotic and nonfluorotic Ae2a,b- and Cftr-null enamel. In hypomineralizing enamel of all models tested, 1) Cl− was strongly reduced; 2) K+ and Na+ accumulated (Na+ not in Amelx-null enamel); and 3) modulation was delayed or blocked. These results suggest that a Na+K+-dependent calcium transporter (likely NCKX4) and a Na+-dependent Pi transporter (potentially NaPi-2b) located in ruffle-ended ameloblasts operate in a coordinated way with the pH-regulating machinery to transport Ca2+, Pi, and bicarbonate into maturation-stage enamel. Acidification and/or associated physicochemical/electrochemical changes in ion levels in enamel fluid near the apical ameloblast membrane may reduce the transport

Trinucleotide Insertions, Deletions, and Point Mutations in Glucose Transporters Confer K+ Uptake in Saccharomyces cerevisiae

PubMed Central

Liang, Hong; Ko, Christopher H.; Herman, Todd; Gaber, Richard F.

1998-01-01

Deletion of TRK1 and TRK2 abolishes high-affinity K+ uptake in Saccharomyces cerevisiae, resulting in the inability to grow on typical synthetic growth medium unless it is supplemented with very high concentrations of potassium. Selection for spontaneous suppressors that restored growth of trk1Δ trk2Δ cells on K+-limiting medium led to the isolation of cells with unusual gain-of-function mutations in the glucose transporter genes HXT1 and HXT3 and the glucose/galactose transporter gene GAL2. 86Rb uptake assays demonstrated that the suppressor mutations conferred increased uptake of the ion. In addition to K+, the mutant hexose transporters also conferred permeation of other cations, including Na+. Because the selection strategy required such gain of function, mutations that disrupted transporter maturation or localization to the plasma membrane were avoided. Thus, the importance of specific sites in glucose transport could be independently assessed by testing for the ability of the mutant transporter to restore glucose-dependent growth to cells containing null alleles of all of the known functional glucose transporter genes. Twelve sites, most of which are conserved among eukaryotic hexose transporters, were revealed to be essential for glucose transport. Four of these have previously been shown to be essential for glucose transport by animal or plant transporters. Eight represented sites not previously known to be crucial for glucose uptake. Each suppressor mutant harbored a single mutation that altered an amino acid(s) within or immediately adjacent to a putative transmembrane domain of the transporter. Seven of 38 independent suppressor mutations consisted of in-frame insertions or deletions. The nature of the insertions and deletions revealed a striking DNA template dependency: each insertion generated a trinucleotide repeat, and each deletion involved the removal of a repeated nucleotide sequence. PMID:9447989

Structure-based functional annotation: yeast ymr099c codes for a D-hexose-6-phosphate mutarotase.

PubMed

Graille, Marc; Baltaze, Jean-Pierre; Leulliot, Nicolas; Liger, Dominique; Quevillon-Cheruel, Sophie; van Tilbeurgh, Herman

2006-10-06

Despite the generation of a large amount of sequence information over the last decade, more than 40% of well characterized enzymatic functions still lack associated protein sequences. Assigning protein sequences to documented biochemical functions is an interesting challenge. We illustrate here that structural genomics may be a reasonable approach in addressing these questions. We present the crystal structure of the Saccharomyces cerevisiae YMR099cp, a protein of unknown function. YMR099cp adopts the same fold as galactose mutarotase and shares the same catalytic machinery necessary for the interconversion of the alpha and beta anomers of galactose. The structure revealed the presence in the active site of a sulfate ion attached by an arginine clamp made by the side chain from two strictly conserved arginine residues. This sulfate is ideally positioned to mimic the phosphate group of hexose 6-phosphate. We have subsequently successfully demonstrated that YMR099cp is a hexose-6-phosphate mutarotase with broad substrate specificity. We solved high resolution structures of some substrate enzyme complexes, further confirming our functional hypothesis. The metabolic role of a hexose-6-phosphate mutarotase is discussed. This work illustrates that structural information has been crucial to assign YMR099cp to the orphan EC activity: hexose-phosphate mutarotase.

Glutamate transporter activity promotes enhanced Na+/K+‐ATPase‐mediated extracellular K+ management during neuronal activity

PubMed Central

Larsen, Brian Roland; Holm, Rikke; Vilsen, Bente

2016-01-01

Key points Management of glutamate and K+ in brain extracellular space is of critical importance to neuronal function.The astrocytic α2β2 Na+/K+‐ATPase isoform combination is activated by the K+ transients occurring during neuronal activity.In the present study, we report that glutamate transporter‐mediated astrocytic Na+ transients stimulate the Na+/K+‐ATPase and thus the clearance of extracellular K+.Specifically, the astrocytic α2β1 Na+/K+‐ATPase subunit combination displays an apparent Na+ affinity primed to react to physiological changes in intracellular Na+.Accordingly, we demonstrate a distinct physiological role in K+ management for each of the two astrocytic Na+/K+‐ATPase β‐subunits. Abstract Neuronal activity is associated with transient [K+]o increases. The excess K+ is cleared by surrounding astrocytes, partly by the Na+/K+‐ATPase of which several subunit isoform combinations exist. The astrocytic Na+/K+‐ATPase α2β2 isoform constellation responds directly to increased [K+]o but, in addition, Na+/K+‐ATPase‐mediated K+ clearance could be governed by astrocytic [Na+]i. During most neuronal activity, glutamate is released in the synaptic cleft and is re‐absorbed by astrocytic Na+‐coupled glutamate transporters, thereby elevating [Na+]i. It thus remains unresolved whether the different Na+/K+‐ATPase isoforms are controlled by [K+]o or [Na+]i during neuronal activity. Hippocampal slice recordings of stimulus‐induced [K+]o transients with ion‐sensitive microelectrodes revealed reduced Na+/K+‐ATPase‐mediated K+ management upon parallel inhibition of the glutamate transporter. The apparent intracellular Na+ affinity of isoform constellations involving the astrocytic β2 has remained elusive as a result of inherent expression of β1 in most cell systems, as well as technical challenges involved in measuring intracellular affinity in intact cells. We therefore expressed the different astrocytic isoform constellations in

Mobility of ions, sugar, and water in the cytoplasm of Xenopus oocytes expressing Na+-coupled sugar transporters (SGLT1)

PubMed Central

Zeuthen, Thomas; Zeuthen, Emil; Klaerke, Dan A

2002-01-01

A model was set up to study water transport in membrane proteins expressed in Xenopus oocytes. The model was tested experimentally using human and rabbit Na+-glucose cotransporters (SGLT1), and was used to explain controversies regarding unstirred layer effects. Cotransport of Na+, sugar and water was monitored by two-electrode voltage clamp and online measurements of oocyte volume. The specific resistance of the oocyte cytoplasm was found by means of microelectrodes to be 263 ± 91 Ω cm (s.d., n = 52), or 2.5 times that of Kulori medium, in agreement with reported values of intracellular ion concentrations and diffusion constants. Osmotically induced volume and resistance changes were compatible with a model of the oocyte in which 37 ± 17 % (s.d., n = 66) of the intracellular volume acts as a free solution while the remainder is inert, being occupied by organelles, etc. The model explains the results of several types of experiments: rapid changes in rates of water cotransport induced by changes in clamp voltage followed by osmotic equilibration in sugar-free conditions; volume changes induced by Na+ transport via the ionophore gramicidin; and uphill water transport. Ethanol (0.5 %) induced a marked swelling of the oocytes of about 16 pl s−1. If the specific inhibitor of SGLT1 phlorizin is added from stock solutions in ethanol, the effect of ethanol obfuscates the effects of the inhibitor. We conclude that the transport parameters derived for water cotransport by the SGLT1 can be attributed to the protein residing in the plasma membrane with no significant influences from unstirred layer effects. PMID:12096052

Continuous production of ethanol from hexoses and pentoses using immobilized mixed cultures of Escherichia coli strains

PubMed Central

Unrean, Pornkamol; Srienc, Friedrich

2010-01-01

We have developed highly efficient ethanologenic E. coli strains that selectively consume pentoses and/or hexoses. Mixed cultures of these strains can be used to selectively adjust the sugar utilization kinetics in ethanol fermentations. Based on the kinetics of sugar utilization, we have designed and implemented an immobilized cell system for the optimized continuous conversion of sugars into ethanol. The results confirm that immobilized mixed cultures support a simultaneous conversion of hexoses and pentoses into ethanol at high yield and at a faster rate than immobilized homogenous cells. Continuous ethanol production has been maintained for several weeks at high productivity with near complete sugar utilization. The control of sugar utilization using immobilized mixed cultures can be adapted to any composition of hexoses and pentoses by adjusting the strain distribution of immobilized cells. The approach, therefore, holds promise for ethanol fermentation from lignocellulosic hydrolysates where the feedstock varies in sugar composition. PMID:20699108

Quantum transport in coupled Majorana box systems

NASA Astrophysics Data System (ADS)

Gau, Matthias; Plugge, Stephan; Egger, Reinhold

2018-05-01

We present a theoretical analysis of low-energy quantum transport in coupled Majorana box devices. A single Majorana box represents a Coulomb-blockaded mesoscopic superconductor proximitizing two or more long topological nanowires. The box thus harbors at least four Majorana zero modes (MZMs). Setups with several Majorana boxes, where MZMs on different boxes are tunnel coupled via short nanowire segments, are key ingredients to recent Majorana qubit and code network proposals. We construct and study the low-energy theory for multiterminal junctions with normal leads connected to the coupled box device by lead-MZM tunnel contacts. Transport experiments in such setups can test the nonlocality of Majorana-based systems and the integrity of the underlying Majorana qubits. For a single box, we recover the previously described topological Kondo effect which can be captured by a purely bosonic theory. For several coupled boxes, however, nonconserved local fermion parities require the inclusion of additional local sets of Pauli operators. We present a renormalization group analysis and develop a nonperturbative strong-coupling approach to quantum transport in such systems. Our findings are illustrated for several examples, including a loop qubit device and different two-box setups.

Phylogenetic evidence for a fusion of archaeal and bacterial SemiSWEETs to form eukaryotic SWEETs and identification of SWEET hexose transporters in the amphibian chytrid pathogen Batrachochytrium dendrobatidis.

PubMed

Hu, Yi-Bing; Sosso, Davide; Qu, Xiao-Qing; Chen, Li-Qing; Ma, Lai; Chermak, Diane; Zhang, De-Chun; Frommer, Wolf B

2016-10-01

SWEETs represent a new class of sugar transporters first described in plants, animals, and humans and later in prokaryotes. Plant SWEETs play key roles in phloem loading, seed filling, and nectar secretion, whereas the role of archaeal, bacterial, and animal transporters remains elusive. Structural analyses show that eukaryotic SWEETs are composed of 2 triple-helix bundles (THBs) fused via an inversion linker helix, whereas prokaryotic SemiSWEETs contain only a single THB and require homodimerization to form transport pores. This study indicates that SWEETs retained sugar transport activity in all kingdoms of life, and that SemiSWEETs are likely their ancestral units. Fusion of oligomeric subunits into single polypeptides during evolution of eukaryotes is commonly found for transporters. Phylogenetic analyses indicate that THBs of eukaryotic SWEETs may not have evolved by tandem duplication of an open reading frame, but rather originated by fusion between an archaeal and a bacterial SemiSWEET, which potentially explains the asymmetry of eukaryotic SWEETs. Moreover, despite the ancient ancestry, SWEETs had not been identified in fungi or oomycetes. Here, we report the identification of SWEETs in oomycetes as well as SWEETs and a potential SemiSWEET in primitive fungi. BdSWEET1 and BdSWEET2 from Batrachochytrium dendrobatidis, a nonhyphal zoosporic fungus that causes global decline in amphibians, showed glucose and fructose transport activities.-Hu, Y.-B., Sosso, D., Qu, X.-Q., Chen, L.-Q., Ma, L., Chermak, D., Zhang, D.-C., Frommer, W. B. Phylogenetic evidence for a fusion of archaeal and bacterial SemiSWEETs to form eukaryotic SWEETs and identification of SWEET hexose transporters in the amphibian chytrid pathogen Batrachochytrium dendrobatidis. © FASEB.

Interface-facilitated energy transport in coupled Frenkel-Kontorova chains

NASA Astrophysics Data System (ADS)

Su, Rui-Xia; Yuan, Zong-Qiang; Wang, Jun; Zheng, Zhi-Gang

2016-04-01

The role of interface couplings on the energy transport of two coupled Frenkel-Kontorova (FK) chains is explored through numerical simulations. In general, it is expected that the interface couplings result in the suppression of heat conduction through the coupled system due to the additional interface phonon-phonon scattering. In the present paper, it is found that the thermal conductivity increases with increasing intensity of interface interactions for weak inter-chain couplings, whereas the heat conduction is suppressed by the interface interaction in the case of strong inter-chain couplings. Based on the phonon spectral energy density method, we demonstrate that the enhancement of energy transport results from the excited phonon modes (in addition to the intrinsic phonon modes), while the strong interface phonon-phonon scattering results in the suppressed energy transport.

The Vitis vinifera sugar transporter gene family: phylogenetic overview and macroarray expression profiling

PubMed Central

2010-01-01

Background In higher plants, sugars are not only nutrients but also important signal molecules. They are distributed through the plant via sugar transporters, which are involved not only in sugar long-distance transport via the loading and the unloading of the conducting complex, but also in sugar allocation into source and sink cells. The availability of the recently released grapevine genome sequence offers the opportunity to identify sucrose and monosaccharide transporter gene families in a woody species and to compare them with those of the herbaceous Arabidopsis thaliana using a phylogenetic analysis. Results In grapevine, one of the most economically important fruit crop in the world, it appeared that sucrose and monosaccharide transporter genes are present in 4 and 59 loci, respectively and that the monosaccharide transporter family can be divided into 7 subfamilies. Phylogenetic analysis of protein sequences has indicated that orthologs exist between Vitis and Arabidospis. A search for cis-regulatory elements in the promoter sequences of the most characterized transporter gene families (sucrose, hexoses and polyols transporters), has revealed that some of them might probably be regulated by sugars. To profile several genes simultaneously, we created a macroarray bearing cDNA fragments specific to 20 sugar transporter genes. This macroarray analysis has revealed that two hexose (VvHT1, VvHT3), one polyol (VvPMT5) and one sucrose (VvSUC27) transporter genes, are highly expressed in most vegetative organs. The expression of one hexose transporter (VvHT2) and two tonoplastic monosaccharide transporter (VvTMT1, VvTMT2) genes are regulated during berry development. Finally, three putative hexose transporter genes show a preferential organ specificity being highly expressed in seeds (VvHT3, VvHT5), in roots (VvHT2) or in mature leaves (VvHT5). Conclusions This study provides an exhaustive survey of sugar transporter genes in Vitis vinifera and revealed that sugar

Models for coupling of salt and water transport; Proximal tubular reabsorption in Necturus kidney.

PubMed

Sackin, H; Boulpaep, E L

1975-12-01

Models for coupling of salt and water transport are developed with two important assumptions appropriate for leaky epithelia. (a) The tight junction is permeable to both sale and water. (b) Active Na transport into the lateral speces is assumed to occur uniformly along the length of the channel. The proposed models deal specifically with the intraepithelial mechanism of proximal tubular resbsorption in the Necturus kidney although they have implications for epithelial transport in the gallbladder and small intestine as well. The first model (continuous version) is similar to the standing gradient model devised by Diamond and Bossert but used different boundary conditions. In contrast to Diamond and Bossert's model, the predicted concentration profiles are relatively flat with no sizable gradients along the interspace. The second model (compartment version) expands Curran's model of epithelial salt and water transport by including additional compartments and considering both electrical and chemical driving forces for individual Na and Cl ions as well as hydraulic and osmotic driving forces for water. In both models, ion and water fluxes are investigated as a function of the transport parameters. The behavior of the models is consistent with previously suggested mechanisms for the control of net transport, particularly during saline diuresis. Under all conditions the predicted ratio of net solute to solvent flux, or emergent concentration, deviates from exact isotonicity (except when the basement membrane has an appreciable salt reflection coefficient). However, the degree of hypertonicity may be small enough to be experimentally indistinguishable from isotonic transport.

Models for coupling of salt and water transport; Proximal tubular reabsorption in Necturus kidney

PubMed Central

Sackin, H; Boulpaep, EL

1975-01-01

Models for coupling of salt and water transport are developed with two important assumptions appropriate for leaky epithelia. (a) The tight junction is permeable to both sale and water. (b) Active Na transport into the lateral speces is assumed to occur uniformly along the length of the channel. The proposed models deal specifically with the intraepithelial mechanism of proximal tubular resbsorption in the Necturus kidney although they have implications for epithelial transport in the gallbladder and small intestine as well. The first model (continuous version) is similar to the standing gradient model devised by Diamond and Bossert but used different boundary conditions. In contrast to Diamond and Bossert's model, the predicted concentration profiles are relatively flat with no sizable gradients along the interspace. The second model (compartment version) expands Curran's model of epithelial salt and water transport by including additional compartments and considering both electrical and chemical driving forces for individual Na and Cl ions as well as hydraulic and osmotic driving forces for water. In both models, ion and water fluxes are investigated as a function of the transport parameters. The behavior of the models is consistent with previously suggested mechanisms for the control of net transport, particularly during saline diuresis. Under all conditions the predicted ratio of net solute to solvent flux, or emergent concentration, deviates from exact isotonicity (except when the basement membrane has an appreciable salt reflection coefficient). However, the degree of hypertonicity may be small enough to be experimentally indistinguishable from isotonic transport. PMID:1104761

Na+-independent D-glucose transport in rabbit renal basolateral membranes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cheung, P.T.; Hammerman, M.R.

1988-05-01

To define the mechanism by which glucose is transported across the basolateral membrane of the renal proximal tubular cell, we measured D-(14C)glucose uptake in basolateral membrane vesicles from rabbit kidney. Na+-dependent D-glucose transport, demonstrable in brush-border vesicles, could not be demonstrated in basolateral membrane vesicles. In the absence of Na+, the uptake of D-(14C)glucose in basolateral vesicles was more rapid than that of L-(3H)glucose over a concentration range of 1-50 mM. Subtraction of the latter from the former uptakes revealed a saturable process with apparent Km of 9.9 mM and Vmax of 0.80 nmol.mg protein-1.s-1. To characterize the transport componentmore » of D-glucose uptake in basolateral vesicles, we measured trans stimulation of 2 mM D-(14C)glucose entry in the absence of Na+. Trans stimulation could be effected by preloading basolateral vesicles with D-glucose, 2-deoxy-D-glucose, or 3-O-methyl-D-glucose, but not with L-glucose or alpha-methyl-D-glucoside. Trans-stimulated D-(14C)glucose uptake was inhibited by 0.1 mM phloretin or cytochalasin B but not phlorizin. In contrast, Na+-dependent D-(14C)glucose transport in brush-border vesicles was inhibited by phlorizin but not phloretin or cytochalasin B. Our findings are consistent with the presence of a Na+-independent D-glucose transporter in the proximal tubular basolateral membrane with characteristics similar to those of transporters present in nonepithelial cells.« less

Shoot Na+ exclusion and increased salinity tolerance engineered by cell type-specific alteration of Na+ transport in Arabidopsis.

PubMed

Møller, Inge S; Gilliham, Matthew; Jha, Deepa; Mayo, Gwenda M; Roy, Stuart J; Coates, Juliet C; Haseloff, Jim; Tester, Mark

2009-07-01

Soil salinity affects large areas of cultivated land, causing significant reductions in crop yield globally. The Na+ toxicity of many crop plants is correlated with overaccumulation of Na+ in the shoot. We have previously suggested that the engineering of Na+ exclusion from the shoot could be achieved through an alteration of plasma membrane Na+ transport processes in the root, if these alterations were cell type specific. Here, it is shown that expression of the Na+ transporter HKT1;1 in the mature root stele of Arabidopsis thaliana decreases Na+ accumulation in the shoot by 37 to 64%. The expression of HKT1;1 specifically in the mature root stele is achieved using an enhancer trap expression system for specific and strong overexpression. The effect in the shoot is caused by the increased influx, mediated by HKT1;1, of Na+ into stelar root cells, which is demonstrated in planta and leads to a reduction of root-to-shoot transfer of Na+. Plants with reduced shoot Na+ also have increased salinity tolerance. By contrast, plants constitutively expressing HKT1;1 driven by the cauliflower mosaic virus 35S promoter accumulated high shoot Na+ and grew poorly. Our results demonstrate that the modification of a specific Na+ transport process in specific cell types can reduce shoot Na+ accumulation, an important component of salinity tolerance of many higher plants.

Comparison of stream reach scale transport of rhodamine WT and NaCl in coupled mountain stream-hyporheic system

Treesearch

A. Bouchier; M. N. Gooseff; B. McGlynn; R. A. Payn; M. A. Briggs

2006-01-01

Rather than interpret the late-time behavior of rhodamine WT (RWT) breakthrough curves in stream tracer studies as indications of hyporheic exchanges we suggest that RWT is lost by sorption. We assessed the transport of RWT compared with NaCl during and after a 5 hour co-injection steady state drip experiment conducted in a headwater mountain stream. We hypothesize...

Transport in Halobacterium Halobium: Light-Induced Cation-Gradients, Amino Acid Transport Kinetics, and Properties of Transport Carriers

NASA Technical Reports Server (NTRS)

Lanyi, Janos K.

1977-01-01

Cell envelope vesicles prepared from H. halobium contain bacteriorhodopsin and upon illumination protons are ejected. Coupled to the proton motive force is the efflux of Na(+). Measurements of Na-22 flux, exterior pH change, and membrane potential, Delta(psi) (with the dye 3,3'-dipentyloxadicarbocyanine) indicate that the means of Na(+) transport is sodium/proton exchange. The kinetics of the pH changes and other evidence suggests that the antiport is electrogenic (H(+)/Na(++ greater than 1). The resulting large chemical gradient for Na(+) (outside much greater than inside), as well as the membrane potential, will drive the transport of 18 amino acids. The I9th, glutamate, is unique in that its accumulation is indifferent to Delta(psi): this amino acid is transported only when a chemical gradient for Na(+) is present. Thus, when more and more NaCl is included in the vesicles glutamate transport proceeds with longer and longer lags. After illumination the gradient of H+() collapses within 1 min, while the large Na(+) gradient and glutamate transporting activity persists for 10- 15 min, indicating that proton motive force is not necessary for transport. A chemical gradient of Na(+), arranged by suspending vesicles loaded with KCl in NaCl, drives glutamate transport in the dark without other sources of energy, with V(sub max) and K(sub m) comparable to light-induced transport. These and other lines of evidence suggest that the transport of glutamate is facilitated by symport with Na(+), in an electrically neutral fashion, so that only the chemical component of the Na(+) gradient is a driving force.

Blocking hexose entry into glycolysis activates alternative metabolic conversion of these sugars and upregulates pentose metabolism in Aspergillus nidulans

DOE PAGES

Khosravi, Claire; Battaglia, Evy; Kun, Roland S.; ...

2018-03-22

Background: Plant biomass is the most abundant carbon source for many fungal species. In the biobased industry fungi are used to produce lignocellulolytic enzymes to degrade agricultural waste biomass. Here we evaluated if it would be possible to create an Aspergillus nidulans strain that releases but does not metabolize hexoses from plant biomass. For this purpose, metabolic mutants were generated that were impaired in glycolysis, by using hexokinase (hxkA) and glucokinase (glkA) negative strains. To prevent repression of enzyme production due to the hexose accumulation, strains were generated that combined these mutations with a deletion in creA, the repressor involvedmore » in regulating preferential use of different carbon catabolic pathways. Results: Phenotypic analysis revealed reduced growth for the hxkA1 glkA4 mutant on wheat bran. However, hexoses did not accumulate during growth of the mutants on wheat bran, suggesting that glucose metabolism is re-routed towards alternative carbon catabolic pathways. The creAΔ4 mutation in combination with preventing initial phosphorylation in glycolysis resulted in better growth than the hxkA/glkA mutant and an increased expression of pentose catabolic and pentose phosphate pathway genes. This indicates that the reduced ability to use hexoses as carbon sources created a shift towards the pentose fraction of wheat bran as a major carbon source to support growth. Conclusion: Blocking the direct entry of hexoses to glycolysis activates alternative metabolic conversion of these sugars in A. nidulans during growth on plant biomass, but also upregulates conversion of other sugars, such as pentoses.« less

Blocking hexose entry into glycolysis activates alternative metabolic conversion of these sugars and upregulates pentose metabolism in Aspergillus nidulans

DOE Office of Scientific and Technical Information (OSTI.GOV)

Khosravi, Claire; Battaglia, Evy; Kun, Roland S.

Background: Plant biomass is the most abundant carbon source for many fungal species. In the biobased industry fungi are used to produce lignocellulolytic enzymes to degrade agricultural waste biomass. Here we evaluated if it would be possible to create an Aspergillus nidulans strain that releases but does not metabolize hexoses from plant biomass. For this purpose, metabolic mutants were generated that were impaired in glycolysis, by using hexokinase (hxkA) and glucokinase (glkA) negative strains. To prevent repression of enzyme production due to the hexose accumulation, strains were generated that combined these mutations with a deletion in creA, the repressor involvedmore » in regulating preferential use of different carbon catabolic pathways. Results: Phenotypic analysis revealed reduced growth for the hxkA1 glkA4 mutant on wheat bran. However, hexoses did not accumulate during growth of the mutants on wheat bran, suggesting that glucose metabolism is re-routed towards alternative carbon catabolic pathways. The creAΔ4 mutation in combination with preventing initial phosphorylation in glycolysis resulted in better growth than the hxkA/glkA mutant and an increased expression of pentose catabolic and pentose phosphate pathway genes. This indicates that the reduced ability to use hexoses as carbon sources created a shift towards the pentose fraction of wheat bran as a major carbon source to support growth. Conclusion: Blocking the direct entry of hexoses to glycolysis activates alternative metabolic conversion of these sugars in A. nidulans during growth on plant biomass, but also upregulates conversion of other sugars, such as pentoses.« less

Fluid dilution and efficiency of Na(+) transport in a mathematical model of a thick ascending limb cell.

PubMed

Nieves-González, Aniel; Clausen, Chris; Marcano, Mariano; Layton, Anita T; Layton, Harold E; Moore, Leon C

2013-03-15

Thick ascending limb (TAL) cells are capable of reducing tubular fluid Na(+) concentration to as low as ~25 mM, and yet they are thought to transport Na(+) efficiently owing to passive paracellular Na(+) absorption. Transport efficiency in the TAL is of particular importance in the outer medulla where O(2) availability is limited by low blood flow. We used a mathematical model of a TAL cell to estimate the efficiency of Na(+) transport and to examine how tubular dilution and cell volume regulation influence transport efficiency. The TAL cell model represents 13 major solutes and the associated transporters and channels; model equations are based on mass conservation and electroneutrality constraints. We analyzed TAL transport in cells with conditions relevant to the inner stripe of the outer medulla, the cortico-medullary junction, and the distal cortical TAL. At each location Na(+) transport efficiency was computed as functions of changes in luminal NaCl concentration ([NaCl]), [K(+)], [NH(4)(+)], junctional Na(+) permeability, and apical K(+) permeability. Na(+) transport efficiency was calculated as the ratio of total net Na(+) transport to transcellular Na(+) transport. Transport efficiency is predicted to be highest at the cortico-medullary boundary where the transepithelial Na(+) gradient is the smallest. Transport efficiency is lowest in the cortex where luminal [NaCl] approaches static head.

SWEETs, transporters for intracellular and intercellular sugar translocation.

PubMed

Eom, Joon-Seob; Chen, Li-Qing; Sosso, Davide; Julius, Benjamin T; Lin, I W; Qu, Xiao-Qing; Braun, David M; Frommer, Wolf B

2015-06-01

Three families of transporters have been identified as key players in intercellular transport of sugars: MSTs (monosaccharide transporters), SUTs (sucrose transporters) and SWEETs (hexose and sucrose transporters). MSTs and SUTs fall into the major facilitator superfamily; SWEETs constitute a structurally different class of transporters with only seven transmembrane spanning domains. The predicted topology of SWEETs is supported by crystal structures of bacterial homologs (SemiSWEETs). On average, angiosperm genomes contain ∼20 paralogs, most of which serve distinct physiological roles. In Arabidopsis, AtSWEET8 and 13 feed the pollen; SWEET11 and 12 provide sucrose to the SUTs for phloem loading; AtSWEET11, 12 and 15 have distinct roles in seed filling; AtSWEET16 and 17 are vacuolar hexose transporters; and SWEET9 is essential for nectar secretion. The remaining family members await characterization, and could play roles in the gametophyte as well as other important roles in sugar transport in the plant. In rice and cassava, and possibly other systems, sucrose transporting SWEETs play central roles in pathogen resistance. Notably, the human genome also contains a glucose transporting isoform. Further analysis promises new insights into mechanism and regulation of assimilate allocation and a new potential for increasing crop yield. Copyright © 2015 Elsevier Ltd. All rights reserved.

NEUTRALIZATION OF THE ASPARTIC ACID RESIDUE D367, BUT NOT D454, INHIBITS BINDING OF NA+ TO THE GLUTAMATE-FREE FORM AND CYCLING OF THE GLUTAMATE TRANSPORTER EAAC1

PubMed Central

Tao, Zhen; Zhang, Zhou; Grewer, Christof

2008-01-01

Substrate transport by the plasma membrane glutamate transporter EAAC1 is coupled to cotransport of three sodium ions. One of these Na+ ions binds to the transporter already in the absence of glutamate. Here, we have investigated the possible involvement of two conserved aspartic acid residues in transmembrane segments 7 and 8 of EAAC1, D367 and D454, in Na+ cotransport. In order to test the effect of charge neutralization mutations in these positions on Na+ binding to the glutamate-free transporter, we recorded the Na+-induced anion leak current to determine the Km of EAAC1 for Na+. For EAAC1WT, this Km was determined as 120 mM. When the negative charge of D367 was neutralized by mutagenesis to asparagine, Na+ activated the anion leak current with a Km of about 2 M, indicating dramatically impaired Na+ binding to the mutant transporter. In contrast, the Na+ affinity of EAAC1D454N was virtually unchanged compared to the wild type transporter (Km = 90 mM). The reduced occupancy of the Na+ binding site of EAAC1D367N resulted in a dramatic reduction in glutamate affinity (Km = 3.6 mM, 140 mM [Na+]), which could be partially overcome by increasing extracellular [Na+]. In addition to impairing Na+ binding, the D367N mutation slowed glutamate transport, as shown by pre-steady-state kinetic analysis of transport currents, by strongly decreasing the rate of a reaction step associated with glutamate translocation. Our data are consistent with a model in which D367, but not D454 is involved in coordinating the bound Na+ in the glutamate-free transporter form. PMID:16478724

Isotonic transport by the Na+-glucose cotransporter SGLT1 from humans and rabbit

PubMed Central

Zeuthen, T; Meinild, A-K; Loo, D D F; Wright, E M; Klaerke, D A

2001-01-01

In order to study its role in steady state water transport, the Na+-glucose cotransporter (SGLT1) was expressed in Xenopus laevis oocytes; both the human and the rabbit clones were tested. The transport activity was monitored as a clamp current and the flux of water followed optically as the change in oocyte volume. SGLT1 has two modes of water transport. First, it acts as a molecular water pump: for each 2 Na+ and 1 sugar molecule 264 water molecules were cotransported in the human SGLT1 (hSGLT1), 424 for the rabbit SGLT1 (rSGLT1). Second, it acts as a water channel. The cotransport of water was tightly coupled to the sugar-induced clamp current. Instantaneous changes in clamp current induced by changes in clamp voltage were accompanied by instantaneous changes in the rate of water transport. The cotransported solution was predicted to be hypertonic, and an osmotic gradient built up across the oocyte membrane with continued transport; this resulted in an additional osmotic influx of water. After 5-10 min a steady state was achieved in which the total influx was predicted to be isotonic with the intracellular solution. With the given expression levels, the steady state water transport was divided about equally between cotransport, osmosis across the SGLT1 and osmosis across the native oocyte membrane. Coexpression of AQP1 with the SGLT1 increased the water permeability more than 10-fold and steady state isotonic transport was achieved after less than 2 s of sugar activation. One-third of the water was cotransported, and the remainder was osmotically driven through the AQP1. The data suggest that SGLT1 has three roles in isotonic water transport: it cotransports water directly, it supplies a passive pathway for osmotic water transport, and it generates an osmotic driving force that can be employed by other pathways, for example aquaporins. PMID:11251046

Coupled transport in field-reversed configurations

NASA Astrophysics Data System (ADS)

Steinhauer, L. C.; Berk, H. L.; TAE Team

2018-02-01

Coupled transport is the close interconnection between the cross-field and parallel fluxes in different regions due to topological changes in the magnetic field. This occurs because perpendicular transport is necessary for particles or energy to leave closed field-line regions, while parallel transport strongly affects evolution of open field-line regions. In most toroidal confinement systems, the periphery, namely, the portion with open magnetic surfaces, is small in thickness and volume compared to the core plasma, the portion with closed surfaces. In field-reversed configurations (FRCs), the periphery plays an outsized role in overall confinement. This effect is addressed by an FRC-relevant model of coupled particle transport that is well suited for immediate interpretation of experiments. The focus here is particle confinement rather than energy confinement since the two track together in FRCs. The interpretive tool yields both the particle transport rate χn and the end-loss time τǁ. The results indicate that particle confinement depends on both χn across magnetic surfaces throughout the plasma and τǁ along open surfaces and that they provide roughly equal transport barriers, inhibiting particle loss. The interpretation of traditional FRCs shows Bohm-like χn and inertial (free-streaming) τǁ. However, in recent advanced beam-driven FRC experiments, χn approaches the classical rate and τǁ is comparable to classic empty-loss-cone mirrors.

The Role of Na+ and K+ Transporters in Salt Stress Adaptation in Glycophytes

PubMed Central

Assaha, Dekoum V. M.; Ueda, Akihiro; Saneoka, Hirofumi; Al-Yahyai, Rashid; Yaish, Mahmoud W.

2017-01-01

Ionic stress is one of the most important components of salinity and is brought about by excess Na+ accumulation, especially in the aerial parts of plants. Since Na+ interferes with K+ homeostasis, and especially given its involvement in numerous metabolic processes, maintaining a balanced cytosolic Na+/K+ ratio has become a key salinity tolerance mechanism. Achieving this homeostatic balance requires the activity of Na+ and K+ transporters and/or channels. The mechanism of Na+ and K+ uptake and translocation in glycophytes and halophytes is essentially the same, but glycophytes are more susceptible to ionic stress than halophytes. The transport mechanisms involve Na+ and/or K+ transporters and channels as well as non-selective cation channels. Thus, the question arises of whether the difference in salt tolerance between glycophytes and halophytes could be the result of differences in the proteins or in the expression of genes coding the transporters. The aim of this review is to seek answers to this question by examining the role of major Na+ and K+ transporters and channels in Na+ and K+ uptake, translocation and intracellular homeostasis in glycophytes. It turns out that these transporters and channels are equally important for the adaptation of glycophytes as they are for halophytes, but differential gene expression, structural differences in the proteins (single nucleotide substitutions, impacting affinity) and post-translational modifications (phosphorylation) account for the differences in their activity and hence the differences in tolerance between the two groups. Furthermore, lack of the ability to maintain stable plasma membrane (PM) potentials following Na+-induced depolarization is also crucial for salt stress tolerance. This stable membrane potential is sustained by the activity of Na+/H+ antiporters such as SOS1 at the PM. Moreover, novel regulators of Na+ and K+ transport pathways including the Nax1 and Nax2 loci regulation of SOS1 expression and

Proton movement and coupling in the POT family of peptide transporters

PubMed Central

Parker, Joanne L.; Li, Chenghan; Brinth, Allete; Wang, Zhi; Vogeley, Lutz; Solcan, Nicolae; Ledderboge-Vucinic, Gregory; Swanson, Jessica M. J.; Caffrey, Martin; Voth, Gregory A.

2017-01-01

POT transporters represent an evolutionarily well-conserved family of proton-coupled transport systems in biology. An unusual feature of the family is their ability to couple the transport of chemically diverse ligands to an inwardly directed proton electrochemical gradient. For example, in mammals, fungi, and bacteria they are predominantly peptide transporters, whereas in plants the family has diverged to recognize nitrate, plant defense compounds, and hormones. Although recent structural and biochemical studies have identified conserved sites of proton binding, the mechanism through which transport is coupled to proton movement remains enigmatic. Here we show that different POT transporters operate through distinct proton-coupled mechanisms through changes in the extracellular gate. A high-resolution crystal structure reveals the presence of ordered water molecules within the peptide binding site. Multiscale molecular dynamics simulations confirm proton transport occurs through these waters via Grotthuss shuttling and reveal that proton binding to the extracellular side of the transporter facilitates a reorientation from an inward- to outward-facing state. Together these results demonstrate that within the POT family multiple mechanisms of proton coupling have likely evolved in conjunction with variation of the extracellular gate. PMID:29180426

Proton movement and coupling in the POT family of peptide transporters.

PubMed

Parker, Joanne L; Li, Chenghan; Brinth, Allete; Wang, Zhi; Vogeley, Lutz; Solcan, Nicolae; Ledderboge-Vucinic, Gregory; Swanson, Jessica M J; Caffrey, Martin; Voth, Gregory A; Newstead, Simon

2017-12-12

POT transporters represent an evolutionarily well-conserved family of proton-coupled transport systems in biology. An unusual feature of the family is their ability to couple the transport of chemically diverse ligands to an inwardly directed proton electrochemical gradient. For example, in mammals, fungi, and bacteria they are predominantly peptide transporters, whereas in plants the family has diverged to recognize nitrate, plant defense compounds, and hormones. Although recent structural and biochemical studies have identified conserved sites of proton binding, the mechanism through which transport is coupled to proton movement remains enigmatic. Here we show that different POT transporters operate through distinct proton-coupled mechanisms through changes in the extracellular gate. A high-resolution crystal structure reveals the presence of ordered water molecules within the peptide binding site. Multiscale molecular dynamics simulations confirm proton transport occurs through these waters via Grotthuss shuttling and reveal that proton binding to the extracellular side of the transporter facilitates a reorientation from an inward- to outward-facing state. Together these results demonstrate that within the POT family multiple mechanisms of proton coupling have likely evolved in conjunction with variation of the extracellular gate. Copyright © 2017 the Author(s). Published by PNAS.

Erythrocyte metabolism in hyperthyroidism: a microcalorimetric study on changes in the Embden-Meyerhof and the hexose monophosphate pathways.

PubMed

Monti, M; Hedner, P; Ikomi-Kumm, J; Valdemarsson, S

1987-05-01

Erythrocyte metabolism was studied in vitro by microcalorimetry in 10 hyperthyroid subjects before and after treatment. By inhibiting the enzyme enolase in the Embden-Meyerhof pathway with sodium fluoride (NaF) we have recorded the anaerobic and aerobic contributions in erythrocyte thermogenesis. The decrease in heat production rate in samples with NaF corresponds to the anaerobic contribution, whereas the values from samples with NaF reflect aerobic processes. Before treatment, total heat production rate was 120 +/- 2 mW/l erythrocytes which was higher than the post-treatment value of 99 +/- 2 (P less than 0.001) as well as the value for 14 euthyroid subjects, 108 +/- 2 mW/l (P less than 0.001). The NaF inhibitable rate was 73 +/- 2 before and 63 +/- 1 mW/l after therapy (P less than 0.01). These values correspond to 61 +/- 1 and 64 +/- 1% (n.s.) of the total heat production rate, and were similar to that of 61 +/- 2% for the controls. Heat production rates in the presence of NaF were 47 +/- 1 before and 36 +/- 1 mW/l after therapy (P less than 0.001), representing 39 +/- 1 and 36 +/- 1% of total values, respectively. The present results show that overall metabolism is increased in erythrocytes from hyperthyroid subjects before treatment and returns to normal after normalization of the thyroid function. Moreover, by using microcalorimetry we found that the metabolic activity along the Embden-Meyerhof anaerobic pathway as well as along the hexose monophosphate aerobic pathway in erythrocytes is stimulated by thyroid hormones.

The Na(+) transporter, TaHKT1;5-D, limits shoot Na(+) accumulation in bread wheat.

PubMed

Byrt, Caitlin Siobhan; Xu, Bo; Krishnan, Mahima; Lightfoot, Damien James; Athman, Asmini; Jacobs, Andrew Keith; Watson-Haigh, Nathan S; Plett, Darren; Munns, Rana; Tester, Mark; Gilliham, Matthew

2014-11-01

Bread wheat (Triticum aestivum L.) has a major salt tolerance locus, Kna1, responsible for the maintenance of a high cytosolic K(+) /Na(+) ratio in the leaves of salt stressed plants. The Kna1 locus encompasses a large DNA fragment, the distal 14% of chromosome 4DL. Limited recombination has been observed at this locus making it difficult to map genetically and identify the causal gene. Here, we decipher the function of TaHKT1;5-D, a candidate gene underlying the Kna1 locus. Transport studies using the heterologous expression systems Saccharomyces cerevisiae and Xenopus laevis oocytes indicated that TaHKT1;5-D is a Na(+) -selective transporter. Transient expression in Arabidopsis thaliana mesophyll protoplasts and in situ polymerase chain reaction indicated that TaHKT1;5-D is localised on the plasma membrane in the wheat root stele. RNA interference-induced silencing decreased the expression of TaHKT1;5-D in transgenic bread wheat lines which led to an increase in the Na(+) concentration in the leaves. This indicates that TaHKT1;5-D retrieves Na(+) from the xylem vessels in the root and has an important role in restricting the transport of Na(+) from the root to the leaves in bread wheat. Thus, TaHKT1;5-D confers the essential salinity tolerance mechanism in bread wheat associated with the Kna1 locus via shoot Na(+) exclusion and is critical in maintaining a high K(+) /Na(+) ratio in the leaves. These findings show there is potential to increase the salinity tolerance of bread wheat by manipulation of HKT1;5 genes. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

Ion-Transport Design for High-Performance Na+-Based Electrochromics.

PubMed

Li, Ran; Li, Kerui; Wang, Gang; Li, Lei; Zhang, Qiangqiang; Yan, Jinhui; Chen, Yao; Zhang, Qinghong; Hou, Chengyi; Li, Yaogang; Wang, Hongzhi

2018-04-24

Sodium ion (Na + )-based electrochemical systems have been extensively investigated in batteries and supercapacitors and also can be quality candidates for electrochromic (EC) devices. However, poor diffusion kinetics and severe EC performance degradation occur during the intercalation/deintercalation processes because the ionic radii of Na + are larger than those of conventional intercalation ions. Here, through intentional design of ion-transport channels in metal-organic frameworks (MOFs), Na + serves as an efficient intercalation ion for incorporation into a nanostructured electrode with a high diffusion coefficient of approximately 10 -8 cm 2 s -1 . As a result, the well-designed MOF-based EC device demonstrates desirable Na + EC performance, including fast switching speed, multicolor switching, and high stability. A smart "quick response code" display is fabricated using a mask-free laser writing method for application in the "Internet of Things". In addition, the concept of ion transport pathway design can be widely adopted for fabricating high-performance ion intercalation materials and devices for consumer electronics.

Tributyltin sensitivity of vacuolar-type Na(+)-transporting ATPase from Enterococcus hirae.

PubMed

Chardwiriyapreecha, Soracom; Inoue, Tomohiro; Sugimoto, Naoko; Sekito, Takayuki; Yamato, Ichiro; Murata, Takeshi; Homma, Michio; Kakinuma, Yoshimi

2009-10-01

Tributyltin chloride (TBT), an environmental pollutant, is toxic to a variety of eukaryotic and prokaryotic organisms. Some members of F-ATP synthase (F-ATPase)/vacuolar type ATPase (V-ATPase) superfamily have been identified as the molecular target of this compound. TBT inhibited the activities of H(+)-transporting or Na(+)-transporting F-ATPase as well as H(+)-transporting V-ATPase originated from various organisms. However, the sensitivity to TBT of Na(+)-transporting V-ATPase has not been investigated. We examined the effect of TBT on Na(+)-transporting V-ATPase from an eubacterium Enterococus hirae. The ATP hydrolytic activity of E. hirae V-ATPase in purified form as well as in membrane-bound form was little inhibited by less than 10 microM TBT; IC50 for TBT inhibition of purified enzyme was estimated to be about 35 microM. Active sodium transport by E. hirae cells, indicating the in vivo activity of this V-ATPase, was not inhibited by 20 microM TBT. By contrast, IC50 of H(+)-transporting V-ATPase of the vacuolar membrane vesicles from Saccharomyces cerevisiae was about 0.2 microM. E. hirae V-ATPase is thus extremely less sensitive to TBT.

Role of PDZK1 Protein in Apical Membrane Expression of Renal Sodium-coupled Phosphate Transporters*

PubMed Central

Giral, Hector; Lanzano, Luca; Caldas, Yupanqui; Blaine, Judith; Verlander, Jill W.; Lei, Tim; Gratton, Enrico; Levi, Moshe

2011-01-01

The sodium-dependent phosphate (Na/Pi) transporters NaPi-2a and NaPi-2c play a major role in the renal reabsorption of Pi. The functional need for several transporters accomplishing the same role is still not clear. However, the fact that these transporters show differential regulation under dietary and hormonal stimuli suggests different roles in Pi reabsorption. The pathways controlling this differential regulation are still unknown, but one of the candidates involved is the NHERF family of scaffolding PDZ proteins. We propose that differences in the molecular interaction with PDZ proteins are related with the differential adaptation of Na/Pi transporters. Pdzk1−/− mice adapted to chronic low Pi diets showed an increased expression of NaPi-2a protein in the apical membrane of proximal tubules but impaired up-regulation of NaPi-2c. These results suggest an important role for PDZK1 in the stabilization of NaPi-2c in the apical membrane. We studied the specific protein-protein interactions of Na/Pi transporters with NHERF-1 and PDZK1 by FRET. FRET measurements showed a much stronger interaction of NHERF-1 with NaPi-2a than with NaPi-2c. However, both Na/Pi transporters showed similar FRET efficiencies with PDZK1. Interestingly, in cells adapted to low Pi concentrations, there were increases in NaPi-2c/PDZK1 and NaPi-2a/NHERF-1 interactions. The differential affinity of the Na/Pi transporters for NHERF-1 and PDZK1 proteins could partially explain their differential regulation and/or stability in the apical membrane. In this regard, direct interaction between NaPi-2c and PDZK1 seems to play an important role in the physiological regulation of NaPi-2c. PMID:21388960

Shoot Na+ Exclusion and Increased Salinity Tolerance Engineered by Cell Type–Specific Alteration of Na+ Transport in Arabidopsis[W][OA

PubMed Central

Møller, Inge S.; Gilliham, Matthew; Jha, Deepa; Mayo, Gwenda M.; Roy, Stuart J.; Coates, Juliet C.; Haseloff, Jim; Tester, Mark

2009-01-01

Soil salinity affects large areas of cultivated land, causing significant reductions in crop yield globally. The Na+ toxicity of many crop plants is correlated with overaccumulation of Na+ in the shoot. We have previously suggested that the engineering of Na+ exclusion from the shoot could be achieved through an alteration of plasma membrane Na+ transport processes in the root, if these alterations were cell type specific. Here, it is shown that expression of the Na+ transporter HKT1;1 in the mature root stele of Arabidopsis thaliana decreases Na+ accumulation in the shoot by 37 to 64%. The expression of HKT1;1 specifically in the mature root stele is achieved using an enhancer trap expression system for specific and strong overexpression. The effect in the shoot is caused by the increased influx, mediated by HKT1;1, of Na+ into stelar root cells, which is demonstrated in planta and leads to a reduction of root-to-shoot transfer of Na+. Plants with reduced shoot Na+ also have increased salinity tolerance. By contrast, plants constitutively expressing HKT1;1 driven by the cauliflower mosaic virus 35S promoter accumulated high shoot Na+ and grew poorly. Our results demonstrate that the modification of a specific Na+ transport process in specific cell types can reduce shoot Na+ accumulation, an important component of salinity tolerance of many higher plants. PMID:19584143

Nerve growth factor reduces amiloride‐sensitive Na+ transport in human airway epithelial cells

PubMed Central

Shimko, Michael J.; Zaccone, Eric J.; Thompson, Janet A.; Schwegler‐Berry, Diane; Kashon, Michael L.; Fedan, Jeffrey S.

2014-01-01

Abstract Nerve growth factor (NGF) is overexpressed in patients with inflammatory lung diseases, including virus infections. Airway surface liquid (ASL), which is regulated by epithelial cell ion transport, is essential for normal lung function. No information is available regarding the effect of NGF on ion transport of airway epithelium. To investigate whether NGF can affect ion transport, human primary air‐interface cultured epithelial cells were placed in Ussing chambers to obtain transepithelial voltage (−7.1 ± 3.4 mV), short‐circuit current (Isc, 5.9 ± 1.0 μA), and transepithelial resistance (750 Ω·cm2), and to measure responses to ion transport inhibitors. Amiloride (apical, 3.5 × 10−5 mol/L) decreased Isc by 55.3%. Apically applied NGF (1 ng/mL) reduced Isc by 5.3% in 5 min; basolaterally applied NGF had no effect. The response to amiloride was reduced (41.6%) in the presence of NGF. K‐252a (10 nmol/L, apical) did not itself affect Na+ transport, but it attenuated the NGF‐induced reduction in Na+ transport, indicating the participation of the trkA receptor in the NGF‐induced reduction in Na+ transport. PD‐98059 (30 μmol/L, apical and basolateral) did not itself affect Na+ transport, but attenuated the NGF‐induced reduction in Na+ transport, indicating that trkA activated the Erk 1/2 signaling cascade. NGF stimulated phosphorylation of Erk 1/2 and the β‐subunit of ENaC. K‐252a and PD‐98059 inhibited these responses. NGF had no effect on Isc in the presence of apical nystatin (50 μmol/L). These results indicate that NGF inhibits Na+ transport through a trkA‐Erk 1/2‐activated signaling pathway linked to ENaC phosphorylation. PMID:25347857

Transport properties of elastically coupled fractional Brownian motors

NASA Astrophysics Data System (ADS)

Lv, Wangyong; Wang, Huiqi; Lin, Lifeng; Wang, Fei; Zhong, Suchuan

2015-11-01

Under the background of anomalous diffusion, which is characterized by the sub-linear or super-linear mean-square displacement in time, we proposed the coupled fractional Brownian motors, in which the asymmetrical periodic potential as ratchet is coupled mutually with elastic springs, and the driving source is the external harmonic force and internal thermal fluctuations. The transport mechanism of coupled particles in the overdamped limit is investigated as the function of the temperature of baths, coupling constant and natural length of the spring, the amplitude and frequency of driving force, and the asymmetry of ratchet potential by numerical stimulations. The results indicate that the damping force involving the information of historical velocity leads to the nonlocal memory property and blocks the traditional dissipative motion behaviors, and it even plays a cooperative role of driving force in drift motion of the coupled particles. Thus, we observe various non-monotonic resonance-like behaviors of collective directed transport in the mediums with different diffusion exponents.

(13)C metabolic flux analysis in neurons utilizing a model that accounts for hexose phosphate recycling within the pentose phosphate pathway.

PubMed

Gebril, Hoda M; Avula, Bharathi; Wang, Yan-Hong; Khan, Ikhlas A; Jekabsons, Mika B

2016-02-01

Glycolysis, mitochondrial substrate oxidation, and the pentose phosphate pathway (PPP) are critical for neuronal bioenergetics and oxidation-reduction homeostasis, but quantitating their fluxes remains challenging, especially when processes such as hexose phosphate (i.e., glucose/fructose-6-phosphate) recycling in the PPP are considered. A hexose phosphate recycling model was developed which exploited the rates of glucose consumption, lactate production, and mitochondrial respiration to infer fluxes through the major glucose consuming pathways of adherent cerebellar granule neurons by replicating [(13)C]lactate labeling from metabolism of [1,2-(13)C2]glucose. Flux calculations were predicated on a steady-state system with reactions having known stoichiometries and carbon atom transitions. Non-oxidative PPP activity and consequent hexose phosphate recycling, as well as pyruvate production by cytoplasmic malic enzyme, were optimized by the model and found to account for 28 ± 2% and 7.7 ± 0.2% of hexose phosphate and pyruvate labeling, respectively. From the resulting fluxes, 52 ± 6% of glucose was metabolized by glycolysis, compared to 19 ± 2% by the combined oxidative/non-oxidative pentose cycle that allows for hexose phosphate recycling, and 29 ± 8% by the combined oxidative PPP/de novo nucleotide synthesis reactions. By extension, 62 ± 6% of glucose was converted to pyruvate, the metabolism of which resulted in 16 ± 1% of glucose oxidized by mitochondria and 46 ± 6% exported as lactate. The results indicate a surprisingly high proportion of glucose utilized by the pentose cycle and the reactions synthesizing nucleotides, and exported as lactate. While the in vitro conditions to which the neurons were exposed (high glucose, no lactate or other exogenous substrates) limit extrapolating these results to the in vivo state, the approach provides a means of assessing a number of metabolic fluxes within the context of hexose phosphate recycling in the PPP from a

Electrophysiology of sodium-coupled transport in proximal renal tubules.

PubMed

Lang, F; Messner, G; Rehwald, W

1986-06-01

Effects of sodium-coupled transport on intracellular electrolytes and electrical properties of proximal renal tubule cells are described in this review. Simultaneous with addition of substrate for sodium-coupled transport to luminal perfusates, both cell membranes depolarize. The luminal cell membrane depolarizes due to opening of sodium-cotransport pathways. The depolarization of the peritubular cell membrane during sodium-coupled transport is primarily due to a circular current reentering the lumen via the paracellular pathway. The depolarization leads to a transient decrease of basolateral potassium conductance that in turn amplifies the depolarization. However, within 5-10 min of continued exposure to substrate, potassium conductance increases again, and peritubular cell membrane repolarizes. During depolarization the driving force of peritubular bicarbonate exit is reduced. As a result net alkalinization of the cell prevails despite an increase of intracellular sodium activity, which reduces the driving force for the sodium-hydrogen ion exchanger and would thus have been expected to acidify the cell. No evidence is obtained for regulatory inhibition of sodium-coupled transport by intracellular sodium or calcium. Rather, luminal cotransport is altered by the change of driving forces.

Insights on Na(+) binding and conformational dynamics in multidrug and toxic compound extrusion transporter NorM.

PubMed

Song, Jianing; Ji, Changge; Zhang, John Z H

2014-02-01

MATE (multidrug and toxic compound extrusion) transporter proteins mediate metabolite transport in plants and multidrug resistance in bacteria and mammals. MATE transporter NorM from Vibrio cholerae is an antiporter that is driven by Na+ gradient to extrude the substrates. To understand the molecular mechanism of Na+-substrate exchange, molecular dynamics simulation was performed to study conformational changes of both wild-type and mutant NorM with and without cation bindings. Our results show that NorM is able to bind two Na(+) ions simultaneously, one to each of the carboxylic groups of E255 and D371 in the binding pocket. Furthermore, this di-Na(+) binding state is likely more efficient for conformational changes of NorM_VC toward the inward-facing conformation than single-Na(+) binding state. The observation of two Na(+) binding sites of NorM_VC is consistent with the previous study that two sites for ion binding (denoted as Na1/Na2 sites) are found in the transporter LeuT and BetP, another two secondary transporters. Taken together, our findings shed light on the structure rearrangements of NorM on Na(+) binding and enrich our knowledge of the transport mechanism of secondary transporters. Copyright © 2013 Wiley Periodicals, Inc.

Functional expression and characterization of the Trypanosoma brucei procyclic glucose transporter, THT2.

PubMed

Barrett, M P; Tetaud, E; Seyfang, A; Bringaud, F; Baltz, T

1995-12-15

The gene encoding THT2, one of two hexose-transporter isoforms present in Trypanosoma brucei, has been expressed in both Xenopus laevis oocytes and a stably transfected line of Chinese hamster ovary (CHO) cells. The heterologously expressed gene encodes a protein with pharmacological and kinetic parameters similar to those of the hexose transporter measured in procyclic-culture-form trypanosomes. The substrate recognition of the THT2 transporter differed from that of the THT1 isoform, which is expressed only in bloodstream forms, in that: (i) it has a relatively high affinity for substrate with a Km of 59 microM for 2-deoxy-D-glucose (2-DOG) and a similar high affinity for D-glucose (compared with Km of 0.5 mM for 2-DOG in bloodstream forms); (ii) the affinity for 6-deoxy-D-glucose (6-DOG) is two orders of magnitude lower than that for D-glucose, whereas the bloodstream-form transporter recognizes D-glucose and its 6-DOG analogue with similar affinity; (iii) the bloodstream-form transporter, but not THT2, recognizes 3-fluoro-3-deoxy-D-glucose. D-Fructose-transport capacity and insensitivity to D-galactose was also found in THT2-expressing CHO cells and procyclic trypanosomes. We conclude from these cumulative results that the THT2 gene encodes the transporter responsible for hexose transport in procyclic trypanosomes. The transport of 2-DOG in procyclic organisms was inhibited by both the protonophore, carbonyl cyanide 4-trifluoromethoxy phenylhydrazone (FCCP), and KCN, suggesting a requirement for a protonmotive force. However, sensitivity to these reagents depended on the external substrate concentration, with uptake being unaffected at substrate concentrations higher than 2 mM. THT2 expressed in CHO cells behaved as a facilitated transporter, and was unaffected by FCCP or KCN over the whole substrate concentration range tested.

The Divergence, Actions, Roles, and Relatives of Sodium-Coupled Bicarbonate Transporters

PubMed Central

Boron, Walter F.

2013-01-01

The mammalian Slc4 (Solute carrier 4) family of transporters is a functionally diverse group of 10 multi-spanning membrane proteins that includes three Cl-HCO3 exchangers (AE1–3), five Na+-coupled HCO3− transporters (NCBTs), and two other unusual members (AE4, BTR1). In this review, we mainly focus on the five mammalian NCBTs-NBCe1, NBCe2, NBCn1, NDCBE, and NBCn2. Each plays a specialized role in maintaining intracellular pH and, by contributing to the movement of HCO3− across epithelia, in maintaining whole-body pH and otherwise contributing to epithelial transport. Disruptions involving NCBT genes are linked to blindness, deafness, proximal renal tubular acidosis, mental retardation, and epilepsy. We also review AE1–3, AE4, and BTR1, addressing their relevance to the study of NCBTs. This review draws together recent advances in our understanding of the phylogenetic origins and physiological relevance of NCBTs and their progenitors. Underlying these advances is progress in such diverse disciplines as physiology, molecular biology, genetics, immunocytochemistry, proteomics, and structural biology. This review highlights the key similarities and differences between individual NCBTs and the genes that encode them and also clarifies the sometimes confusing NCBT nomenclature. PMID:23589833

100-W 105-μm 0.15NA fiber coupled laser diode module

NASA Astrophysics Data System (ADS)

Karlsen, Scott R.; Price, R. Kirk; Reynolds, Mitch; Brown, Aaron; Mehl, Ron; Patterson, Steve; Martinsen, Robert J.

2009-02-01

We report on the development of a high brightness laser diode module capable of coupling over 100W of optical power into a 105 μm 0.15 NA fiber at 976 nm. This module, based on nLIGHT's Pearl product architecture, utilizes hard soldered single emitters packaged into a compact and passively-cooled package. In this system each diode is individually collimated in the fast and slow axes and free-space coupled into a single fiber. The high brightness module has an optical excitation under 0.13 NA, is virtually free of cladding modes, and has an electrical to optical efficiency greater than 40%. Additionally, this module is compatible with high power 7:1 fused fiber combiners, and initial experiments demonstrated 500W coupled into a 220 μm, 0.22 NA fiber. These modules address the need in the market for higher brightness diode lasers for pumping fiber lasers and direct material processing.

Mechanism of epithelial lithium transport. Evidence for basolateral Na:Na and Na:Li exchange

PubMed Central

1983-01-01

Measurement of transmural sodium fluxes across isolated, ouabain- inhibited turtle colon in the presence of a serosal-to-mucosal sodium gradient shows that in the absence of active transport the amiloride- sensitive cellular path contains at least two routes for the transmural movement of sodium and lithium, one a conductive path and the other a nonconductive, cation-exchange mechanism. The latter transport element can exchange lithium for sodium, and the countertransport of these two cations provides a mechanistic basis for the ability of tight epithelia to actively absorb lithium despite the low affinity of the basolateral Na/K-ATPase for this cation. PMID:6644269

Reduced Cellular Mg2+ Content Enhances Hexose 6-Phosphate Dehydrogenase Activity and Expression in HepG2 and HL-60 Cells

PubMed Central

Voma, Chesinta; Barfell, Andrew; Croniger, Colleen; Romani, Andrea

2014-01-01

We have reported that Mg2+ dynamically regulates glucose 6-phosphate entry into the endoplasmic reticulum and its hydrolysis by the glucose 6-phosphatase in liver cells. In the present study, we report that by modulating glucose 6-phosphate entry into the endoplasmic reticulum of HepG2 cells, Mg2+ also regulates the oxidation of this substrate via hexose 6-phosphate dehydrogenase (H6PD). This regulatory effect is dynamic as glucose 6-phosphate entry and oxidation can be rapidly down-regulated by the addition of exogenous Mg2+. In addition, HepG2 cells growing in low Mg2+ show a marked increase in hexose 6-phosphate dehydrogenase mRNA and protein expression. Metabolically, these effects on hexose 6-phosphate dehydrogenase are important as this enzyme increases intra-reticular NADPH production, which favors fatty acid and cholesterol synthesis. Similar effects of Mg2+ were observed in HL-60 cells. These and previously published results suggest that in an hepatocyte culture model changes in cytoplasmic Mg2+ content regulates glucose 6-phosphate utilization via glucose 6 phosphatase and hexose-6 phosphate dehydrogenase in alternative to glycolysis and glycogen synthesis. This alternative regulation might be of relevance in the transition from fed to fasted state. PMID:24631573

Coupled Neutron Transport for HZETRN

NASA Technical Reports Server (NTRS)

Slaba, Tony C.; Blattnig, Steve R.

2009-01-01

Exposure estimates inside space vehicles, surface habitats, and high altitude aircrafts exposed to space radiation are highly influenced by secondary neutron production. The deterministic transport code HZETRN has been identified as a reliable and efficient tool for such studies, but improvements to the underlying transport models and numerical methods are still necessary. In this paper, the forward-backward (FB) and directionally coupled forward-backward (DC) neutron transport models are derived, numerical methods for the FB model are reviewed, and a computationally efficient numerical solution is presented for the DC model. Both models are compared to the Monte Carlo codes HETC-HEDS, FLUKA, and MCNPX, and the DC model is shown to agree closely with the Monte Carlo results. Finally, it is found in the development of either model that the decoupling of low energy neutrons from the light particle transport procedure adversely affects low energy light ion fluence spectra and exposure quantities. A first order correction is presented to resolve the problem, and it is shown to be both accurate and efficient.

23Na and 39K NMR studies of ion transport in human erythrocytes.

PubMed

Ogino, T; Shulman, G I; Avison, M J; Gullans, S R; den Hollander, J A; Shulman, R G

1985-02-01

Ion transport in human erythrocytes was studied by 23Na and 39K NMR with an anionic paramagnetic shift reagent, Dy(P3O10)2(7-). The intra- and extracellular 23Na and 39K NMR signals were well separated (over 10 ppm) at 5 mM concentration of the shift reagent. The NMR visibility of the intracellular Na+ and K+ was determined to be 100% in human and duck erythrocytes. The intracellular ion concentrations were 8.1 +/- 0.8 mM Na+ (n = 7) and 110 +/- 12 mM K+ (n = 4) for fresh human erythrocytes. The ouabain-sensitive net Na+ efflux was 1.75 +/- 0.08 mmol/hr per liter of cells at 37 degrees C (n = 3). The gramicidin-induced ion transport in human erythrocytes was also studied by 23Na and 39K NMR or by simultaneous measurements of 23Na NMR and a K+-selective electrode. The time courses of the Na+ and K+ transport induced by the ionophore were biphasic. The initial rapid fluxes were due to an exchange of Na+ for K+, which were found to occur with a 1:1 stoichiometry. The subsequent slow components were the net Na+ and K+ effluxes rate-limited by the Cl- permeability and accompanied by a reduction in cell volume. The Cl- permeability determined from the NMR measurements of these slow fluxes was 3.2 +/- 0.5 X 10(-8) cm/sec at 25 degrees C (n = 4).

23Na and 39K NMR studies of ion transport in human erythrocytes.

PubMed Central

Ogino, T; Shulman, G I; Avison, M J; Gullans, S R; den Hollander, J A; Shulman, R G

1985-01-01

Ion transport in human erythrocytes was studied by 23Na and 39K NMR with an anionic paramagnetic shift reagent, Dy(P3O10)2(7-). The intra- and extracellular 23Na and 39K NMR signals were well separated (over 10 ppm) at 5 mM concentration of the shift reagent. The NMR visibility of the intracellular Na+ and K+ was determined to be 100% in human and duck erythrocytes. The intracellular ion concentrations were 8.1 +/- 0.8 mM Na+ (n = 7) and 110 +/- 12 mM K+ (n = 4) for fresh human erythrocytes. The ouabain-sensitive net Na+ efflux was 1.75 +/- 0.08 mmol/hr per liter of cells at 37 degrees C (n = 3). The gramicidin-induced ion transport in human erythrocytes was also studied by 23Na and 39K NMR or by simultaneous measurements of 23Na NMR and a K+-selective electrode. The time courses of the Na+ and K+ transport induced by the ionophore were biphasic. The initial rapid fluxes were due to an exchange of Na+ for K+, which were found to occur with a 1:1 stoichiometry. The subsequent slow components were the net Na+ and K+ effluxes rate-limited by the Cl- permeability and accompanied by a reduction in cell volume. The Cl- permeability determined from the NMR measurements of these slow fluxes was 3.2 +/- 0.5 X 10(-8) cm/sec at 25 degrees C (n = 4). PMID:2579385

Na(+)-dependent transport of taurine is found only on the abluminal membrane of the blood-brain barrier.

PubMed

Rasgado-Flores, Hector; Mokashi, Ashwini; Hawkins, Richard A

2012-01-01

Luminal and abluminal plasma membranes were isolated from bovine brain microvessels and used to identify and characterize Na(+)-dependent and facilitative taurine transport. The calculated transmembrane potential was -59 mV at time 0; external Na(+) (or choline under putative zero-trans conditions) was 126 mM (T=25 °C). The apparent affinity constants of the taurine transporters were determined over a range of taurine concentrations from 0.24 μM to 11.4 μM. Abluminal membranes had both Na(+)-dependent taurine transport as well as facilitative transport while luminal membranes only had facilitative transport. The apparent K(m) for facilitative and Na(+)-dependent taurine transport were 0.06±0.02 μM and 0.7±0.1 μM, respectively. The Na(+)-dependent transport of taurine was voltage dependent over the range of voltages studied (-25 to -101 mV). The transport was over 5 times greater at -101 mV compared to when V(m) was -25 mV. The sensitivity to external osmolality of Na(+)-dependent transport was studied over a range of osmolalities (229 to 398 mOsm/kg H(2)O) using mannitol as the osmotic agent to adjust the osmolality. For these experiments the concentration of Na(+) was maintained constant at 50mM, and the calculated transmembrane potential was -59 mV. The Na(+)-dependent transport system was sensitive to osmolality with the greatest rate observed at 229 mOsm/kg H(2)O. Copyright © 2011 Elsevier Inc. All rights reserved.

Coupling between H+ transport and anaerobic glycolysis in turtle urinary bladder: effect of inhibitors of H+ ATPase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Steinmetz, P.R.; Husted, R.F.; Mueller, A.

1981-03-15

The coupling between H+ transport (JH) and anaerobic glycolysis was examined in vitro in an anaerobic preparation of turtle urinary bladder. JH was measured as the short-circuit current after Na+ transport was abolished with ouabain and by pH stat titration. The media were gassed with N2 and 1% CO2 (PO2 less than 0.5 mm Hg) and contained 10 mM glucose. Under these conditions, JH was not inhibited by 3 mM serosal (S) cyanide or by 0.1 mM mucosal (M) dinitrophenol. Control anaerobic lactate production (Jlac) of 47 bladders was plotted as a function of simultaneously measured JH. The slope ofmore » Jlac on JH was 0.58« less

Quantum transport in coupled resonators enclosed synthetic magnetic flux

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jin, L., E-mail: jinliang@nankai.edu.cn

Quantum transport properties are instrumental to understanding quantum coherent transport processes. Potential applications of quantum transport are widespread, in areas ranging from quantum information science to quantum engineering, and not restricted to quantum state transfer, control and manipulation. Here, we study light transport in a ring array of coupled resonators enclosed synthetic magnetic flux. The ring configuration, with an arbitrary number of resonators embedded, forms a two-arm Aharonov–Bohm interferometer. The influence of magnetic flux on light transport is investigated. Tuning the magnetic flux can lead to resonant transmission, while half-integer magnetic flux quantum leads to completely destructive interference and transmissionmore » zeros in an interferometer with two equal arms. -- Highlights: •The light transport is investigated through ring array of coupled resonators enclosed synthetic magnetic field. •Aharonov–Bohm ring interferometer of arbitrary configuration is investigated. •The half-integer magnetic flux quantum leads to destructive interference and transmission zeros for two-arm at equal length. •Complete transmission is available via tuning synthetic magnetic flux.« less

Thermodynamic analysis of active sodium and potassium transport in the frog corneal epithelium.

PubMed

Candia, O A; Reinach, P S

1982-06-01

The formalism of linear nonequilibrium thermodynamics for a three-flow system was applied to the isolated frog corneal epithelium to study the coupling between metabolism and the Na-K transport system across this layer. There is little or no net ion transport across the isolated frog corneal epithelium bathed in Na2SO4 Ringer. Addition of amphotericin B to the tear side solution increases apical membrane permeability, which results in a net Na transport (from tear to stroma) and a net K transport in the opposite direction. Corneas were mounted in a modified Ussing chamber that permitted the simultaneous measurements of electrical parameters and O2 consumption by means of Clark-type oxygen electrodes. The overall degree of coupling, q, of the Na-K transport system to metabolism was calculated from measuring the suprabasal O2 consumption rate at "static head" and "level flow" conditions and by a second independent technique. Measurements of electrical conductance used in conjunction with other previously measured parameters allowed the calculation of the affinity, A, of the metabolic reaction driving transport, all phenomenological coefficients, and the electromotive forces of sodium (ENa) and potassium transport (EK). Values of q determined by the two techniques agreed (q = 0.80 and 0.84, respectively). This indicates incomplete coupling and a variable stoichiometric relationship among O2 consumption rate, net Na transport, and net K transport. The value calculated for A was 70.5 kcal.mol-1, for ENa 142.5 mV, and for EK -34.9 mV.

OsHKT1;4-mediated Na(+) transport in stems contributes to Na(+) exclusion from leaf blades of rice at the reproductive growth stage upon salt stress.

PubMed

Suzuki, Kei; Yamaji, Naoki; Costa, Alex; Okuma, Eiji; Kobayashi, Natsuko I; Kashiwagi, Tatsuhiko; Katsuhara, Maki; Wang, Cun; Tanoi, Keitaro; Murata, Yoshiyuki; Schroeder, Julian I; Ma, Jian Feng; Horie, Tomoaki

2016-01-19

Na(+) exclusion from leaf blades is one of the key mechanisms for glycophytes to cope with salinity stress. Certain class I transporters of the high-affinity K(+) transporter (HKT) family have been demonstrated to mediate leaf blade-Na(+) exclusion upon salinity stress via Na(+)-selective transport. Multiple HKT1 transporters are known to function in rice (Oryza sativa). However, the ion transport function of OsHKT1;4 and its contribution to the Na(+) exclusion mechanism in rice remain to be elucidated. Here, we report results of the functional characterization of the OsHKT1;4 transporter in rice. OsHKT1;4 mediated robust Na(+) transport in Saccharomyces cerevisiae and Xenopus laevis oocytes. Electrophysiological experiments demonstrated that OsHKT1;4 shows strong Na(+) selectivity among cations tested, including Li(+), Na(+), K(+), Rb(+), Cs(+), and NH4 (+), in oocytes. A chimeric protein, EGFP-OsHKT1;4, was found to be functional in oocytes and targeted to the plasma membrane of rice protoplasts. The level of OsHKT1;4 transcripts was prominent in leaf sheaths throughout the growth stages. Unexpectedly however, we demonstrate here accumulation of OsHKT1;4 transcripts in the stem including internode II and peduncle in the reproductive growth stage. Moreover, phenotypic analysis of OsHKT1;4 RNAi plants in the vegetative growth stage revealed no profound influence on the growth and ion accumulation in comparison with WT plants upon salinity stress. However, imposition of salinity stress on the RNAi plants in the reproductive growth stage caused significant Na(+) overaccumulation in aerial organs, in particular, leaf blades and sheaths. In addition, (22)Na(+) tracer experiments using peduncles of RNAi and WT plants suggested xylem Na(+) unloading by OsHKT1;4. Taken together, our results indicate a newly recognized function of OsHKT1;4 in Na(+) exclusion in stems together with leaf sheaths, thus excluding Na(+) from leaf blades of a japonica rice cultivar in the

Coregulated Expression of the Na+/Phosphate Pho89 Transporter and Ena1 Na+-ATPase Allows Their Functional Coupling under High-pH Stress

PubMed Central

Serra-Cardona, Albert; Petrezsélyová, Silvia; Canadell, David; Ramos, José

2014-01-01

The yeast Saccharomyces cerevisiae has two main high-affinity inorganic phosphate (Pi) transporters, Pho84 and Pho89, that are functionally relevant at acidic/neutral pH and alkaline pH, respectively. Upon Pi starvation, PHO84 and PHO89 are induced by the activation of the PHO regulon by the binding of the Pho4 transcription factor to specific promoter sequences. We show that PHO89 and PHO84 are induced by alkalinization of the medium with different kinetics and that the network controlling Pho89 expression in response to alkaline pH differs from that of other members of the PHO regulon. In addition to Pho4, the PHO89 promoter is regulated by the transcriptional activator Crz1 through the calcium-activated phosphatase calcineurin, and it is under the control of several repressors (Mig2, Nrg1, and Nrg2) coordinately regulated by the Snf1 protein kinase and the Rim101 transcription factor. This network mimics the one regulating expression of the Na+-ATPase gene ENA1, encoding a major determinant for Na+ detoxification. Our data highlight a scenario in which the activities of Pho89 and Ena1 are functionally coordinated to sustain growth in an alkaline environment. PMID:25266663

Coupling of active motion and advection shapes intracellular cargo transport.

PubMed

Khuc Trong, Philipp; Guck, Jochen; Goldstein, Raymond E

2012-07-13

Intracellular cargo transport can arise from passive diffusion, active motor-driven transport along cytoskeletal filament networks, and passive advection by fluid flows entrained by such cargo-motor motion. Active and advective transport are thus intrinsically coupled as related, yet different representations of the same underlying network structure. A reaction-advection-diffusion system is used here to show that this coupling affects the transport and localization of a passive tracer in a confined geometry. For sufficiently low diffusion, cargo localization to a target zone is optimized either by low reaction kinetics and decoupling of bound and unbound states, or by a mostly disordered cytoskeletal network with only weak directional bias. These generic results may help to rationalize subtle features of cytoskeletal networks, for example as observed for microtubules in fly oocytes.

Unidirectional Flux Balance of Monovalent Ions in Cells with Na/Na and Li/Na Exchange: Experimental and Computational Studies on Lymphoid U937 Cells

PubMed Central

Vereninov, Igor A.; Yurinskaya, Valentina E.; Model, Michael A.; Vereninov, Alexey A.

2016-01-01

Monovalent ion traffic across the cell membrane occurs via various pathways. Evaluation of individual fluxes in whole cell is hampered by their strong interdependence. This difficulty can be overcome by computational analysis of the whole cell flux balance. However, the previous computational studies disregarded ion movement of the self-exchange type. We have taken this exchange into account. The developed software allows determination of unidirectional fluxes of all monovalent ions via the major pathways both under the balanced state and during transient processes. We show how the problem of finding the rate coefficients can be solved by measurement of monovalent ion concentrations and some of the fluxes. Interdependence of fluxes due to the mandatory conditions of electroneutrality and osmotic balance and due to specific effects can be discriminated, enabling one to identify specific changes in ion transfer machinery under varied conditions. To test the effectiveness of the developed approach we made use of the fact that Li/Na exchange is known to be an analogue of the coupled Na/Na exchange. Thus, we compared the predicted and experimental data obtained on U937 cells under varied Li+ concentrations and following inhibition of the sodium pump with ouabain. We found that the coupled Na/Na exchange in U937 cells comprises a significant portion of the entire Na+ turnover. The data showed that the loading of the sodium pump by Li/Na exchange involved in the secondary active Li+ transport at 1–10 mM external Li+ is small. This result may be extrapolated to similar Li+ and Na+ flux relationships in erythrocytes and other cells in patients treated with Li+ in therapeutic doses. The developed computational approach is applicable for studying various cells and can be useful in education for demonstrating the effects of individual transporters and channels on ion gradients, cell water content and membrane potential. PMID:27159324

Na+/Ca2+ exchange and Na+/K+-ATPase in the heart

PubMed Central

Shattock, Michael J; Ottolia, Michela; Bers, Donald M; Blaustein, Mordecai P; Boguslavskyi, Andrii; Bossuyt, Julie; Bridge, John H B; Chen-Izu, Ye; Clancy, Colleen E; Edwards, Andrew; Goldhaber, Joshua; Kaplan, Jack; Lingrel, Jerry B; Pavlovic, Davor; Philipson, Kenneth; Sipido, Karin R; Xie, Zi-Jian

2015-01-01

This paper is the third in a series of reviews published in this issue resulting from the University of California Davis Cardiovascular Symposium 2014: Systems approach to understanding cardiac excitation–contraction coupling and arrhythmias: Na+ channel and Na+ transport. The goal of the symposium was to bring together experts in the field to discuss points of consensus and controversy on the topic of sodium in the heart. The present review focuses on cardiac Na+/Ca2+ exchange (NCX) and Na+/K+-ATPase (NKA). While the relevance of Ca2+ homeostasis in cardiac function has been extensively investigated, the role of Na+ regulation in shaping heart function is often overlooked. Small changes in the cytoplasmic Na+ content have multiple effects on the heart by influencing intracellular Ca2+ and pH levels thereby modulating heart contractility. Therefore it is essential for heart cells to maintain Na+ homeostasis. Among the proteins that accomplish this task are the Na+/Ca2+ exchanger (NCX) and the Na+/K+ pump (NKA). By transporting three Na+ ions into the cytoplasm in exchange for one Ca2+ moved out, NCX is one of the main Na+ influx mechanisms in cardiomyocytes. Acting in the opposite direction, NKA moves Na+ ions from the cytoplasm to the extracellular space against their gradient by utilizing the energy released from ATP hydrolysis. A fine balance between these two processes controls the net amount of intracellular Na+ and aberrations in either of these two systems can have a large impact on cardiac contractility. Due to the relevant role of these two proteins in Na+ homeostasis, the emphasis of this review is on recent developments regarding the cardiac Na+/Ca2+ exchanger (NCX1) and Na+/K+ pump and the controversies that still persist in the field. PMID:25772291

Sodium ion transport mechanisms in antiperovskite electrolytes Na 3OBr and Na 4OI 2: An in Situ neutron diffraction study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhu, Jinlong; Wang, Yonggang; Li, Shuai

Na-rich antiperovskites are recently developed solid electrolytes with enhanced sodium ionic conductivity and show promising functionality as a novel solid electrolyte in an all solid-stat battery. In this work, the sodium ionic transport pathways of the parent compound Na 3OBr, as well as the modified layered antiperovskite Na 4OI 2, were studied and compared through temperature dependent neutron diffraction combined with the maximum entropy method. In the cubic Na 3OBr antiperovskite, the nuclear density distribution maps at 500 K indicate that sodium ions ho within and among oxygen octahedra, and Br - ions are not involved in the tetragonal Namore » 4OI 2 antiperovskite, Na ions, which connect octahedra in the ab plane, have the lowest activation energy barrier. In conclusion, the transport of sodium ions along the c axis is assisted by I - ions.« less

Sodium ion transport mechanisms in antiperovskite electrolytes Na 3OBr and Na 4OI 2: An in Situ neutron diffraction study

DOE PAGES

Zhu, Jinlong; Wang, Yonggang; Li, Shuai; ...

2016-06-02

Na-rich antiperovskites are recently developed solid electrolytes with enhanced sodium ionic conductivity and show promising functionality as a novel solid electrolyte in an all solid-stat battery. In this work, the sodium ionic transport pathways of the parent compound Na 3OBr, as well as the modified layered antiperovskite Na 4OI 2, were studied and compared through temperature dependent neutron diffraction combined with the maximum entropy method. In the cubic Na 3OBr antiperovskite, the nuclear density distribution maps at 500 K indicate that sodium ions ho within and among oxygen octahedra, and Br - ions are not involved in the tetragonal Namore » 4OI 2 antiperovskite, Na ions, which connect octahedra in the ab plane, have the lowest activation energy barrier. In conclusion, the transport of sodium ions along the c axis is assisted by I - ions.« less

Charge transport in strongly coupled quantum dot solids

NASA Astrophysics Data System (ADS)

Kagan, Cherie R.; Murray, Christopher B.

2015-12-01

The emergence of high-mobility, colloidal semiconductor quantum dot (QD) solids has triggered fundamental studies that map the evolution from carrier hopping through localized quantum-confined states to band-like charge transport in delocalized and hybridized states of strongly coupled QD solids, in analogy with the construction of solids from atoms. Increased coupling in QD solids has led to record-breaking performance in QD devices, such as electronic transistors and circuitry, optoelectronic light-emitting diodes, photovoltaic devices and photodetectors, and thermoelectric devices. Here, we review the advances in synthesis, assembly, ligand treatments and doping that have enabled high-mobility QD solids, as well as the experiments and theory that depict band-like transport in the QD solid state. We also present recent QD devices and discuss future prospects for QD materials and device design.

Charge transport in strongly coupled quantum dot solids.

PubMed

Kagan, Cherie R; Murray, Christopher B

2015-12-01

The emergence of high-mobility, colloidal semiconductor quantum dot (QD) solids has triggered fundamental studies that map the evolution from carrier hopping through localized quantum-confined states to band-like charge transport in delocalized and hybridized states of strongly coupled QD solids, in analogy with the construction of solids from atoms. Increased coupling in QD solids has led to record-breaking performance in QD devices, such as electronic transistors and circuitry, optoelectronic light-emitting diodes, photovoltaic devices and photodetectors, and thermoelectric devices. Here, we review the advances in synthesis, assembly, ligand treatments and doping that have enabled high-mobility QD solids, as well as the experiments and theory that depict band-like transport in the QD solid state. We also present recent QD devices and discuss future prospects for QD materials and device design.

The effect of coupled transport phenomena in the Opalinus Clay and implications for radionuclide transport

NASA Astrophysics Data System (ADS)

Soler, Josep M.

2001-12-01

In this study, the potential effects of coupled transport phenomena on radionuclide transport in the vicinity of a repository for vitrified high-level radioactive waste (HLW) and spent nuclear fuel (SF) hosted by the Opalinus Clay in Switzerland, at times equal to or greater than the expected lifetime of the waste canisters (about 1000 years), are addressed. The solute fluxes associated with advection, chemical diffusion, thermal and chemical osmosis, hyperfiltration and thermal diffusion have been incorporated into a simple one-dimensional transport equation. The analytical solution of this equation, with appropriate parameters, shows that thermal osmosis is the only coupled transport mechanism that could, on its own, have a strong effect on repository performance. Based on the results from the analytical model, two-dimensional finite-difference models incorporating advection and thermal osmosis, and taking conservation of fluid mass into account, have been formulated. The results show that, under the conditions in the vicinity of the repository at the time scales of interest, and due to the constraints imposed by conservation of fluid mass, the advective component of flow will oppose and cancel the thermal-osmotic component. The overall conclusion is that coupled phenomena will only have a very minor impact on radionuclide transport in the Opalinus Clay, in terms of fluid and solute fluxes, at least under the conditions prevailing at times equal to or greater than the expected lifetime of the waste canisters (about 1000 years).

Molecular characteristics of mammalian and insect amino acid transporters: implications for amino acid homeostasis.

PubMed

Castagna, M; Shayakul, C; Trotti, D; Sacchi, V F; Harvey, W R; Hediger, M A

1997-01-01

In mammalian cells, the uptake of amino acids is mediated by specialized, energy-dependent and passive transporters with overlapping substrate specificities. Most energy-dependent transporters are coupled either to the cotransport of Na+ or Cl- or to the countertransport of K+. Passive transporters are either facilitated transporters or channels. As a prelude to the molecular characterization of the different classes of transporters, we have isolated transporter cDNAs by expression-cloning with Xenopus laevis oocytes and we have characterized the cloned transporters functionally by uptake studies into oocytes using radiolabelled substrates and by electrophysiology to determine substrate-evoked currents. Mammalian transporters investigated include the dibasic and neutral amino acid transport protein D2/NBAT (system b0+) and the Na(+)- and K(+)-dependent neuronal and epithelial high-affinity glutamate transporter EAAC1 (system XAG-). A detailed characterization of these proteins has provided new information on transport characteristics and mechanisms for coupling to different inorganic ions. This work has furthermore advanced our understanding of the roles these transporters play in amino acid homeostasis and in various pathologies. For example, in the central nervous system, glutamate transporters are critically important in maintaining the extracellular glutamate concentration below neurotoxic levels, and defects of the human D2 gene have been shown to account for the formation of kidney stones in patients with cystinuria. Using similar approaches, we are investigating the molecular characteristics of K(+)-coupled amino acid transporters in the larval lepidopteran insect midgut. In the larval midgut, K+ is actively secreted into the lumen through the concerted action of an apical H+ V-ATPase and an apical K+/2H+ antiporter, thereby providing the driving force for absorption of amino acids. In vivo, the uptake occurs at extremely high pH (pH 10) and is driven by a

Proton-coupled sugar transport in the prototypical major facilitator superfamily protein XylE

PubMed Central

Wisedchaisri, Goragot; Park, Min-Sun; Iadanza, Matthew G.; Zheng, Hongjin; Gonen, Tamir

2014-01-01

The major facilitator superfamily (MFS) is the largest collection of structurally related membrane proteins that transport a wide array of substrates. The proton-coupled sugar transporter XylE is the first member of the MFS that has been structurally characterized in multiple transporting conformations, including both the outward and inward-facing states. Here we report the crystal structure of XylE in a new inward-facing open conformation, allowing us to visualize the rocker-switch movement of the N-domain against the C-domain during the transport cycle. Using molecular dynamics simulation, and functional transport assays, we describe the movement of XylE that facilitates sugar translocation across a lipid membrane and identify the likely candidate proton-coupling residues as the conserved Asp27 and Arg133. This study addresses the structural basis for proton-coupled substrate transport and release mechanism for the sugar porter family of proteins. PMID:25088546

Route, mechanism, and implications of proton import during Na+/K+ exchange by native Na+/K+-ATPase pumps

PubMed Central

Vedovato, Natascia

2014-01-01

A single Na+/K+-ATPase pumps three Na+ outwards and two K+ inwards by alternately exposing ion-binding sites to opposite sides of the membrane in a conformational sequence coupled to pump autophosphorylation from ATP and auto-dephosphorylation. The larger flow of Na+ than K+ generates outward current across the cell membrane. Less well understood is the ability of Na+/K+ pumps to generate an inward current of protons. Originally noted in pumps deprived of external K+ and Na+ ions, as inward current at negative membrane potentials that becomes amplified when external pH is lowered, this proton current is generally viewed as an artifact of those unnatural conditions. We demonstrate here that this inward current also flows at physiological K+ and Na+ concentrations. We show that protons exploit ready reversibility of conformational changes associated with extracellular Na+ release from phosphorylated Na+/K+ pumps. Reversal of a subset of these transitions allows an extracellular proton to bind an acidic side chain and to be subsequently released to the cytoplasm. This back-step of phosphorylated Na+/K+ pumps that enables proton import is not required for completion of the 3 Na+/2 K+ transport cycle. However, the back-step occurs readily during Na+/K+ transport when external K+ ion binding and occlusion are delayed, and it occurs more frequently when lowered extracellular pH raises the probability of protonation of the externally accessible carboxylate side chain. The proton route passes through the Na+-selective binding site III and is distinct from the principal pathway traversed by the majority of transported Na+ and K+ ions that passes through binding site II. The inferred occurrence of Na+/K+ exchange and H+ import during the same conformational cycle of a single molecule identifies the Na+/K+ pump as a hybrid transporter. Whether Na+/K+ pump–mediated proton inflow may have any physiological or pathophysiological significance remains to be clarified. PMID

Efficient Coupling of Fluid-Plasma and Monte-Carlo-Neutrals Models for Edge Plasma Transport

NASA Astrophysics Data System (ADS)

Dimits, A. M.; Cohen, B. I.; Friedman, A.; Joseph, I.; Lodestro, L. L.; Rensink, M. E.; Rognlien, T. D.; Sjogreen, B.; Stotler, D. P.; Umansky, M. V.

2017-10-01

UEDGE has been valuable for modeling transport in the tokamak edge and scrape-off layer due in part to its efficient fully implicit solution of coupled fluid neutrals and plasma models. We are developing an implicit coupling of the kinetic Monte-Carlo (MC) code DEGAS-2, as the neutrals model component, to the UEDGE plasma component, based on an extension of the Jacobian-free Newton-Krylov (JFNK) method to MC residuals. The coupling components build on the methods and coding already present in UEDGE. For the linear Krylov iterations, a procedure has been developed to ``extract'' a good preconditioner from that of UEDGE. This preconditioner may also be used to greatly accelerate the convergence rate of a relaxed fixed-point iteration, which may provide a useful ``intermediate'' algorithm. The JFNK method also requires calculation of Jacobian-vector products, for which any finite-difference procedure is inaccurate when a MC component is present. A semi-analytical procedure that retains the standard MC accuracy and fully kinetic neutrals physics is therefore being developed. Prepared for US DOE by LLNL under Contract DE-AC52-07NA27344 and LDRD project 15-ERD-059, by PPPL under Contract DE-AC02-09CH11466, and supported in part by the U.S. DOE, OFES.

Transport behaviors of locally fractional coupled Brownian motors with fluctuating interactions

NASA Astrophysics Data System (ADS)

Wang, Huiqi; Ni, Feixiang; Lin, Lifeng; Lv, Wangyong; Zhu, Hongqiang

2018-09-01

In some complex viscoelastic mediums, it is ubiquitous that absorbing and desorbing surrounding Brownian particles randomly occur in coupled systems. The conventional method is to model a variable-mass system driven by both multiplicative and additive noises. In this paper, an improved mathematical model is created based on generalized Langevin equations (GLE) to characterize the random interaction with locally fluctuating number of coupled particles in the elastically coupled factional Brownian motors (FBM). By the numerical simulations, the effect of fluctuating interactions on collective transport behaviors is investigated, and some abnormal phenomena, such as cooperative behaviors, stochastic resonance (SR) and anomalous transport, are observed in the regime of sub-diffusion.

Regulation of sugar transport and metabolism by the Candida albicans Rgt1 transcriptional repressor.

PubMed

Sexton, Jessica A; Brown, Victoria; Johnston, Mark

2007-10-01

The ability of the fungal pathogen Candida albicans to cause systemic infections depends in part on the function of Hgt4, a cell surface sugar sensor. The orthologues of Hgt4 in Saccharomyces cerevisiae, Snf3 and Rgt2, initiate a signalling cascade that inactivates Rgt1, a transcriptional repressor of genes encoding hexose transporters. To determine whether Hgt4 functions similarly through the C. albicans orthologue of Rgt1, we analysed Cargt1 deletion mutants. We found that Cargt1 mutants are sensitive to the glucose analogue 2-deoxyglucose, a phenotype probably due to uncontrolled expression of genes encoding glucose transporters. Indeed, transcriptional profiling revealed that expression of about two dozen genes, including multiple HGT genes encoding hexose transporters, is increased in the Cargt1 mutant in the absence of sugars, suggesting that CaRgt1 represses expression of several HGT genes under this condition. Some of the HGT genes (probably encoding high-affinity transporters) are also repressed by high levels of glucose, and we show that this repression is mediated by CaMig1, the orthologue of the major glucose-activated repressor in S. cerevisiae, but not by its paralogue CaMig2. Therefore, CaRgt1 and CaMig1 collaborate to control expression of C. albicans hexose transporters in response to different levels of sugars. We were surprised to find that CaRgt1 also regulates expression of GAL1, suggesting that regulation of galactose metabolism in C. albicans is unconventional. Finally, Cargt1 mutations cause cells to hyperfilament, and suppress the hypofilamented phenotype of an hgt4 mutant, indicating that the Hgt4 glucose sensor may affect filamentation by modulating sugar import and metabolism via CaRgt1. Copyright 2007 John Wiley & Sons, Ltd.

Critical Roles of Two Hydrophobic Residues within Human Glucose Transporter 9 (hSLC2A9) in Substrate Selectivity and Urate Transport*

PubMed Central

Long, Wentong; Panwar, Pankaj; Witkowska, Kate; Wong, Kenneth; O'Neill, Debbie; Chen, Xing-Zhen; Lemieux, M. Joanne; Cheeseman, Chris I.

2015-01-01

High blood urate levels (hyperuricemia) have been found to be a significant risk factor for cardiovascular diseases and inflammatory arthritis, such as hypertension and gout. Human glucose transporter 9 (hSLC2A9) is an essential protein that mainly regulates urate/hexose homeostasis in human kidney and liver. hSLC2A9 is a high affinity-low capacity hexose transporter and a high capacity urate transporter. Our previous studies identified a single hydrophobic residue in trans-membrane domain 7 of class II glucose transporters as a determinant of fructose transport. A mutation of isoleucine 335 to valine (I355V) in hSLC2A9 can reduce fructose transport while not affecting glucose fluxes. This current study demonstrates that the I335V mutant transports urate similarly to the wild type hSLC2A9; however, Ile-335 is necessary for urate/fructose trans-acceleration exchange to occur. Furthermore, Trp-110 is a critical site for urate transport. Two structural models of the class II glucose transporters, hSLC2A9 and hSLC2A5, based on the crystal structure of hSLC2A1 (GLUT1), reveal that Ile-335 (or the homologous Ile-296 in hSLC2A5) is a key component for protein conformational changes when the protein translocates substrates. The hSLC2A9 model also predicted that Trp-110 is a crucial site that could directly interact with urate during transport. Together, these studies confirm that hSLC2A9 transports both urate and fructose, but it interacts with them in different ways. Therefore, this study advances our understanding of how hSLC2A9 mediates urate and fructose transport, providing further information for developing pharmacological agents to treat hyperuricemia and related diseases, such as gout, hypertension, and diabetes. PMID:25922070

Transport of H(+), Na(+) and K(+) across the posterior midgut of blood-fed mosquitoes (Aedes aegypti).

PubMed

Pacey, Evan K; O'Donnell, Michael J

2014-02-01

Following ingestion of a blood meal, the adult female mosquito undergoes a massive diuresis during which Na(+), Cl(-) and water are secreted at high rates by the Malpighian tubules. In the hours following completion of diuresis, digestion of the K(+)-rich blood cells provides a source of energy as well as amino acids for proteins in the developing eggs. Although the transport of inorganic ions by the Malpighian tubules of blood-fed mosquitoes has been extensively characterized, relatively little is known of the epithelial transport mechanisms responsible for movement of Na(+), H(+), and K(+) across the posterior midgut. In this paper we have used the Scanning Ion-selective Electrode Technique (SIET) to measure the basal (unstimulated) rates of transport of K(+), Na(+) and H(+) across the isolated posterior midgut at intervals after the blood meal. We have also measured luminal concentrations of Na(+) and K(+) and the transepithelial electrical potential at the same time points and have calculated the electrochemical potentials for Na(+), K(+) and H(+) across the midgut. SIET measurements reveal absorption (lumen to bath) of Na(+) and H(+) and secretion of K(+) for the first 2h after blood-feeding. By 24h after the meal, absorption of Na(+) and H(+) remains active while there is an electrochemical gradient favouring absorption of K(+). Inhibition by ouabain and Ba(2+) suggest a role for the Na(+)/K(+)-ATPase and K(+) channels in absorption of Na(+) and K(+), respectively. Inhibition of H(+) absorption by acetazolamide implicates carbonic anhydrase in transepithelial H(+) transport. Copyright © 2014 Elsevier Ltd. All rights reserved.

Visualizing the kinetic power stroke that drives proton-coupled Zn(II) transport

PubMed Central

Gupta, Sayan; Chai, Jin; Cheng, Jie; D'Mello, Rhijuta; Chance, Mark R.; Fu, Dax

2014-01-01

The proton gradient is a principal energy source for respiration-dependent active transport, but the structural mechanisms of proton-coupled transport processes are poorly understood. YiiP is a proton-coupled zinc transporter found in the cytoplasmic membrane of E. coli, and the transport-site of YiiP receives protons from water molecules that gain access to its hydrophobic environment and transduces the energy of an inward proton gradient to drive Zn(II) efflux1,2. This membrane protein is a well characterized member3-7 of the protein family of cation diffusion facilitators (CDFs) that occurs at all phylogenetic levels8-10. X-ray mediated hydroxyl radical labeling of YiiP and mass spectrometric analysis showed that Zn(II) binding triggered a highly localized, all-or-none change of water accessibility to the transport-site and an adjacent hydrophobic gate. Millisecond time-resolved dynamics revealed a concerted and reciprocal pattern of accessibility changes along a transmembrane helix, suggesting a rigid-body helical reorientation linked to Zn(II) binding that triggers the closing of the hydrophobic gate. The gated water access to the transport-site enables a stationary proton gradient to facilitate the conversion of zinc binding energy to the kinetic power stroke of a vectorial zinc transport. The kinetic details provide energetic insights into a proton-coupled active transport reaction. PMID:25043033

The Split Personality of Glutamate Transporters: A Chloride Channel and a Transporter.

PubMed

Cater, Rosemary J; Ryan, Renae M; Vandenberg, Robert J

2016-03-01

Transporters and ion channels are conventionally categorised into distinct classes of membrane proteins. However, some membrane proteins have a split personality and can function as both transporters and ion channels. The excitatory amino acid transporters (EAATs) in particular, function as both glutamate transporters and chloride (Cl(-)) channels. The EAATs couple the transport of glutamate to the co-transport of three Na(+) ions and one H(+) ion into the cell, and the counter-transport of one K(+) ion out of the cell. The EAAT Cl(-) channel is activated by the binding of glutamate and Na(+), but is thermodynamically uncoupled from glutamate transport and involves molecular determinants distinct from those responsible for glutamate transport. Several crystal structures of an EAAT archaeal homologue, GltPh, at different stages of the transport cycle, alongside numerous functional studies and molecular dynamics simulations, have provided extensive insights into the mechanism of substrate transport via these transporters. However, the molecular determinants involved in Cl(-) permeation, and the mechanism by which this channel is activated are not entirely understood. Here we will discuss what is currently known about the molecular determinants involved in EAAT-mediated Cl(-) permeation and the mechanisms that underlie their split personality.

A Loose Relationship: Incomplete H+/Sugar Coupling in the MFS Sugar Transporter GlcP.

PubMed

Bazzone, Andre; Zabadne, Annas J; Salisowski, Anastasia; Madej, M Gregor; Fendler, Klaus

2017-12-19

The glucose transporter from Staphylococcus epidermidis, GlcP Se , is a homolog of the human GLUT sugar transporters of the major facilitator superfamily. Together with the xylose transporter from Escherichia coli, XylE Ec , the other prominent prokaryotic GLUT homolog, GlcP Se , is equipped with a conserved proton-binding site arguing for an electrogenic transport mode. However, the electrophysiological analysis of GlcP Se presented here reveals important differences between the two GLUT homologs. GlcP Se , unlike XylE Ec , does not perform steady-state electrogenic transport at symmetrical pH conditions. Furthermore, when a pH gradient is applied, partially uncoupled transport modes can be generated. In contrast to other bacterial sugar transporters analyzed so far, in GlcP Se sugar binding, translocation and release are also accomplished by the deprotonated transporter. Based on these experimental results, we conclude that coupling of sugar and H + transport is incomplete in GlcP Se . To verify the viability of the observed partially coupled GlcP Se transport modes, we propose a universal eight-state kinetic model in which any degree of coupling is realized and H + /sugar symport represents only a specific instance. Furthermore, using sequence comparison with strictly coupled XylE Ec and similar sugar transporters, we identify an additional charged residue that may be essential for effective H + /sugar symport. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Underexpression of the Na+-dependent neutral amino acid transporter ASCT2 in the spontaneously hypertensive rat kidney.

PubMed

Pinho, Maria João; Pinto, Vanda; Serrão, Maria Paula; Jose, Pedro A; Soares-da-Silva, Patrício

2007-07-01

This study examined the inward transport of l-[(14)C]alanine, an ASCT2 preferential substrate, in monolayers of immortalized renal proximal tubular epithelial (PTE) cells from Wistar-Kyoto (WKY) and spontaneously hypertensive (SHR) rats. The expression of ASCT2 in WKY and SHR PTE cells and kidney cortices from WKY and SHR was also evaluated. l-[(14)C]alanine uptake was highly dependent on extracellular Na(+). Replacement of NaCl by LiCl or choline chloride abolished transport activity in SHR and WKY PTE cells. In the presence of the system L inhibitor BCH, Na(+)-dependent l-alanine uptake in WKY and SHR PTE cells was inhibited by alanine, serine, and cysteine, which is consistent with amino acid transport through ASCT2. The saturable component of Na(+)-dependent l-alanine transport under V(max) conditions in SHR PTE cells was one-half of that in WKY PTE cells, with similar K(m) values. Differences in magnitude of Na(+)-dependent l-alanine uptake through ASCT2 between WKY and SHR PTE cells correlated positively with differences in ASCT2 protein expression, this being more abundant in WKY PTE cells. Abundance of ASCT2 transcript and protein in kidney cortices of SHR rats was also lower than that in normotensive WKY rats. In conclusion, immortalized SHR and WKY PTE cells take up l-alanine mainly through a high-affinity Na(+)-dependent amino acid transporter, with functional features of ASCT2 transport. The activity and expression of the ASCT2 transporter were considerably lower in the SHR cells.

Altered erythrocyte sodium-lithium counter-transport and Na+/K(+)-ATPase activity in cystic fibrosis.

PubMed

Luczay, A; Vásárhelyi, B; Dobos, M; Holics, K; Ujhelyi, R; Tulassay, T

1997-03-01

Patients with cystic fibrosis (CF) exhibit normal concentrations of sodium and chloride in spite of the disturbance of Cl- and Na+ transport in epithelial cells. To characterize compensatory mechanisms in the regulation of sodium homeostasis, erythrocytes of 13 CF patients were analysed for sodium-lithium counter-transport (SLC), Na+/K(+)-ATPase activity and intracellular sodium content. Values were compared to those of healthy controls. Patients with CF had normal serum sodium and chloride concentrations and renal excretions of these ions were within the physiological range. Intracellular sodium concentration was similar in the CF and the control group (6.8 +/- 2.2 vs 5.7 +/- 1.0 mmol/l RBCs). Red blood cells' SLC and Na+/ K(+)-ATPase activity were elevated in CF patients (381 +/- 106 mumol/h/l RBCs vs 281 +/- 64; p < 0.01) and (445 +/- 129 mumol ATP mg prot/h vs 322 +/- 84, p < 0.01). Our study demonstrates that transmembrane cation transport systems are highly activated in CF. The increased sodium transport may be part of a compensatory mechanism of sodium homeostasis in children with CF.

Impact of pore size variability and network coupling on electrokinetic transport in porous media

NASA Astrophysics Data System (ADS)

Alizadeh, Shima; Bazant, Martin Z.; Mani, Ali

2016-11-01

We have developed and validated an efficient and robust computational model to study the coupled fluid and ion transport through electrokinetic porous media, which are exposed to external gradients of pressure, electric potential, and concentration. In our approach a porous media is modeled as a network of many pores through which the transport is described by the coupled Poisson-Nernst-Planck-Stokes equations. When the pore sizes are random, the interactions between various modes of transport may provoke complexities such as concentration polarization shocks and internal flow circulations. These phenomena impact mixing and transport in various systems including deionization and filtration systems, supercapacitors, and lab-on-a-chip devices. In this work, we present simulations of massive networks of pores and we demonstrate the impact of pore size variation, and pore-pore coupling on the overall electrokinetic transport in porous media.

Energetics of Na + Transport through the Electrode/Cathode Interface in Single Solvent Electrolytes

DOE PAGES

Browning, Katie L.; Sacci, Robert L.; Veith, Gabriel M.

2017-01-27

Here, the activation energy of desolvation for Na-ion electrolyte systems were studied using temperature dependent electrochemical impedance spectroscopy (EIS). Propylene carbonate (PC) and tetraglyme were chosen as solvents in single solvent electrolyte solution coupled with NaClO 4, NaPF 6, NaAsF 6, NaBF 4, and NaOFt salts. The results demonstrate the insertion and desolvation processes are extremely fast at or above room temperature. The data shows under optimal salt chemistry the desolvation activation energy is less than 21 kJ/mole. This is in contrast to the ~50 kJ/mole measured for analogous Li systems. The dominant factor affecting performance was the anion. Indeed,more » anions with lower donor numbers (PF 6 –, AsF 6 –) had more than 30 kJ/mole lower desolvation energies than species like NaBF 4 and NaOFt.« less

Mechanism of potassium ion uptake by the Na+/K+-ATPase

PubMed Central

Castillo, Juan P.; Rui, Huan; Basilio, Daniel; Das, Avisek; Roux, Benoît; Latorre, Ramon; Bezanilla, Francisco; Holmgren, Miguel

2015-01-01

The Na+/K+-ATPase restores sodium (Na+) and potassium (K+) electrochemical gradients dissipated by action potentials and ion-coupled transport processes. As ions are transported, they become transiently trapped between intracellular and extracellular gates. Once the external gate opens, three Na+ ions are released, followed by the binding and occlusion of two K+ ions. While the mechanisms of Na+ release have been well characterized by the study of transient Na+ currents, smaller and faster transient currents mediated by external K+ have been more difficult to study. Here we show that external K+ ions travelling to their binding sites sense only a small fraction of the electric field as they rapidly and simultaneously become occluded. Consistent with these results, molecular dynamics simulations of a pump model show a wide water-filled access channel connecting the binding site to the external solution. These results suggest a mechanism of K+ gating different from that of Na+ occlusion. PMID:26205423

A Model to Couple Flow, Thermal and Reactive Chemical Transport, and Geo-mechanics in Variably Saturated Media

NASA Astrophysics Data System (ADS)

Yeh, G. T.; Tsai, C. H.

2015-12-01

This paper presents the development of a THMC (thermal-hydrology-mechanics-chemistry) process model in variably saturated media. The governing equations for variably saturated flow and reactive chemical transport are obtained based on the mass conservation principle of species transport supplemented with Darcy's law, constraint of species concentration, equation of states, and constitutive law of K-S-P (Conductivity-Degree of Saturation-Capillary Pressure). The thermal transport equation is obtained based on the conservation of energy. The geo-mechanic displacement is obtained based on the assumption of equilibrium. Conventionally, these equations have been implicitly coupled via the calculations of secondary variables based on primary variables. The mechanisms of coupling have not been obvious. In this paper, governing equations are explicitly coupled for all primary variables. The coupling is accomplished via the storage coefficients, transporting velocities, and conduction-dispersion-diffusion coefficient tensor; one set each for every primary variable. With this new system of equations, the coupling mechanisms become clear. Physical interpretations of every term in the coupled equations will be discussed. Examples will be employed to demonstrate the intuition and superiority of these explicit coupling approaches. Keywords: Variably Saturated Flow, Thermal Transport, Geo-mechanics, Reactive Transport.

K+ Transport by the OsHKT2;4 Transporter from Rice with Atypical Na+ Transport Properties and Competition in Permeation of K+ over Mg2+ and Ca2+ Ions1[C][W][OA

PubMed Central

Horie, Tomoaki; Brodsky, Dennis E.; Costa, Alex; Kaneko, Toshiyuki; Lo Schiavo, Fiorella; Katsuhara, Maki; Schroeder, Julian I.

2011-01-01

Members of class II of the HKT transporters, which have thus far only been isolated from grasses, were found to mediate Na+-K+ cotransport and at high Na+ concentrations preferred Na+-selective transport, depending on the ionic conditions. But the physiological functions of this K+-transporting class II of HKT transporters remain unknown in plants, with the exception of the unique class II Na+ transporter OsHKT2;1. The genetically tractable rice (Oryza sativa; background Nipponbare) possesses two predicted K+-transporting class II HKT transporter genes, OsHKT2;3 and OsHKT2;4. In this study, we have characterized the ion selectivity of the class II rice HKT transporter OsHKT2;4 in yeast and Xenopus laevis oocytes. OsHKT2;4 rescued the growth defect of a K+ uptake-deficient yeast mutant. Green fluorescent protein-OsHKT2;4 is targeted to the plasma membrane in transgenic plant cells. OsHKT2;4-expressing oocytes exhibited strong K+ permeability. Interestingly, however, K+ influx in OsHKT2;4-expressing oocytes did not require stimulation by extracellular Na+, in contrast to other class II HKT transporters. Furthermore, OsHKT2;4-mediated currents exhibited permeabilities to both Mg2+ and Ca2+ in the absence of competing K+ ions. Comparative analyses of Ca2+ and Mg2+ permeabilities in several HKT transporters, including Arabidopsis thaliana HKT1;1 (AtHKT1;1), Triticum aestivum HKT2;1 (TaHKT2;1), OsHKT2;1, OsHKT2;2, and OsHKT2;4, revealed that only OsHKT2;4 and to a lesser degree TaHKT2;1 mediate Mg2+ transport. Interestingly, cation competition analyses demonstrate that the selectivity of both of these class II HKT transporters for K+ is dominant over divalent cations, suggesting that Mg2+ and Ca2+ transport via OsHKT2;4 may be small and would depend on competing K+ concentrations in plants. PMID:21610181

INTEX-NA: Intercontinental Chemical Transport Experiment - North America

NASA Technical Reports Server (NTRS)

Singh, Hanwant B.; Jacob, D.; Pfister, L.; Hipskind, R. Stephen (Technical Monitor)

2002-01-01

INTEX-NA is an integrated atmospheric chemistry field experiment to be performed over North America using the NASA DC-8 and P-3B aircraft as its primary platforms. It seeks to understand the exchange of chemicals and aerosols between continents and the global troposphere. The constituents of interest are ozone and its precursors (hydrocarbons, NOX and HOX), aerosols, and the major greenhouse gases (CO2, CH4, N2O). INTEX-NA will provide the observational database needed to quantify inflow, outflow, and transformations of chemicals over North America. INTEX-NA is to be performed in two phases. Phase A will take place during the period of May-August 2004 and Phase B during March-June 2006. Phase A is in summer when photochemistry is most intense and climatic issues involving aerosols and carbon cycle are most pressing, and Phase B is in spring when Asian transport to North America is at its peak. INTEX-NA will coordinate its activities with concurrent measurement programs including satellites (e. g. Terra, Aura, Envisat), field activities undertaken by the North American Carbon Program (NACP), and other U.S. and international partners. However, it is being designed as a 'stand alone' mission such that its successful execution is not contingent on other programs. Synthesis of the ensemble of observation from surface, airborne, and space platforms, with the help of global/regional models is an important It is anticipated that approximately 175 flight hours for each of the aircraft (DC-8 and P-3B) will be required for each Phase. Principal operational sites are tentatively selected to be Bangor, ME; Wallops Island, VA; Seattle, WA; Rhinelander, WI; Lancaster, CA; and New Orleans, LA. These coastal and continental sites can support large missions and are suitable for INTEX-NA objectives. The experiment will be supported by forecasts from meteorological and chemical models, satellite observations, surface networks, and enhanced O3,-sonde releases. In addition to

Effects of anion transport inhibitors and ion substitution on Cl sup minus transport in TAL of Henle's loop

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kondo, Yoshiaki; Yoshitomi, Koji; Imai, Masashi

1987-12-01

To identify the mechanism of Cl{sup {minus}} transport across the thin ascending limb of Henle's loop (TAL), the authors examined effects of anion transport inhibitors and ionic substitution in the isolated segments of hamsters using the in vitro microperfusion technique. 4,4{prime}-diisothiocyanostilbene-2,2{prime}-disulfonic acid (DIDS) at 10{sup {minus}3} M changed the NaCl diffusion voltage (V{sub t}) to the orientation that corresponds to the decrease in the Cl{sup {minus}}-Na{sup +} permeability ratio when it was added either to the bath or to the lumen. DIDS, added to the bath or to the lumen decreased the lumen-to-bath flux coefficient for {sup 36}Cl, whereas itmore » had little effect on the flux coefficient for {sup 22}Na. The inhibitory effect of phloretin was rapid and reversible. Phloridzin was ineffective. From these observations, they conclude that Cl{sup {minus}} transport across the TAL is distinct from Na{sup +} and is not coupled with Na{sup +}, K{sup +}, or HCO{sup {minus}}{sub 3}.« less

120W, NA_0.15 fiber coupled LD module with 125-μm clad/NA 0.22 fiber by spatial coupling method

NASA Astrophysics Data System (ADS)

Ishige, Yuta; Kaji, Eisaku; Katayama, Etsuji; Ohki, Yutaka; Gajdátsy, Gábor; Cserteg, András.

2018-02-01

We have fabricated a fiber coupled semiconductor laser diode module by means of spatial beam combining of single emitter broad area semiconductor laser diode chips in the 9xx nm band. In the spatial beam multiplexing method, the numerical aperture of the output light from the optical fiber increases by increasing the number of laser diodes coupled into the fiber. To reduce it, we have tried the approach to improving assembly process technology. As a result, we could fabricate laser diode modules having a light output power of 120W or more and 95% power within NA of 0.15 or less from a single optical fiber with 125-μm cladding diameter. Furthermore, we have obtained that the laser diode module maintaining high coupling efficiency can be realized even around the fill factor of 0.95. This has been achieved by improving the optical alignment method regarding the fast axis stack pitch of the laser diodes in the laser diode module. Therefore, without using techniques such as polarization combining and wavelength combining, high output power was realized while keeping small numerical aperture. This contributes to a reduction in unit price per light output power of the pumping laser diode module.

Single molecule imaging of conformational dynamics in sodium-coupled transporters

NASA Astrophysics Data System (ADS)

Terry, Daniel S.

Neurotransmitter:sodium symporter (NSS) proteins remove neurotransmitters released into the synapse through a transport process driven by the physiological sodium ion (Na+) gradient. NSSs for dopamine, noradrenaline, and serotonin are targeted by the psychostimulants cocaine and amphetamines, as well as by antidepressants. The crystal structure of LeuT, a prokaryotic NSS homologue, revealed the NSS molecular architecture and has been the basis for extensive structural, biochemical, and computational investigations of the mechanism of transporter proteins with a LeuT-like fold. In this dissertation, the conformational states sampled by LeuT are explored using single-molecule fluorescence resonance energy transfer imaging methods, with special focus on the motions of transmembrane helix 1a that lead to inward release of substrate. We also explored how dynamics are modulated by substrate, Na+, and protons to produce efficient transport. These advances represent a first of a kind study of the dynamics of an integral membrane protein at a truly single-molecule scale. Advances in instrumentation, analysis tools, and organic fluorophores were all required to achieve these goals, and such advances are also described. While these experiments were performed with detergent-solubilized protein, preliminary work suggests that imaging of LeuT in proteoliposomes is feasible and a fluorescence sensor assay could be used to simultaneously detect conformational dynamics and transport function.

Single Molecule Imaging of Conformational Dynamics in Sodium-Coupled Transporters

ERIC Educational Resources Information Center

Terry, Daniel S.

2013-01-01

Neurotransmitter:sodium symporter (NSS) proteins remove neurotransmitters released into the synapse through a transport process driven by the physiological sodium ion (Na[superscript +]) gradient. NSSs for dopamine, noradrenaline, and serotonin are targeted by the psychostimulants cocaine and amphetamines, as well as by antidepressants. The…

Antiresonance and decoupling in electronic transport through parallel-coupled quantum-dot structures with laterally-coupled Majorana zero modes

NASA Astrophysics Data System (ADS)

Zhang, Ya-Jing; Zhang, Lian-Lian; Jiang, Cui; Gong, Wei-Jiang

2018-02-01

We theoretically investigate the electronic transport through a parallel-coupled multi-quantum-dot system, in which the terminal dots of a one-dimensional quantum-dot chain are embodied in the two arms of an Aharonov-Bohm interferometer. It is found that in the structures of odd(even) dots, all their even(odd) molecular states have opportunities to decouple from the leads, and in this process antiresonance occurs which are accordant with the odd(even)-numbered eigenenergies of the sub-molecule without terminal dots. Next when Majorana zero modes are introduced to couple laterally to the terminal dots, the antiresonance and decoupling phenomena still co-exist in the quantum transport process. Such a result can be helpful in understanding the special influence of Majorana zero mode on the electronic transport through quantum-dot systems.

Unbiased Simulations Reveal the Inward-Facing Conformation of the Human Serotonin Transporter and Na+ Ion Release

PubMed Central

Koldsø, Heidi; Noer, Pernille; Grouleff, Julie; Autzen, Henriette Elisabeth; Sinning, Steffen; Schiøtt, Birgit

2011-01-01

Monoamine transporters are responsible for termination of synaptic signaling and are involved in depression, control of appetite, and anxiety amongst other neurological processes. Despite extensive efforts, the structures of the monoamine transporters and the transport mechanism of ions and substrates are still largely unknown. Structural knowledge of the human serotonin transporter (hSERT) is much awaited for understanding the mechanistic details of substrate translocation and binding of antidepressants and drugs of abuse. The publication of the crystal structure of the homologous leucine transporter has resulted in homology models of the monoamine transporters. Here we present extended molecular dynamics simulations of an experimentally supported homology model of hSERT with and without the natural substrate yielding a total of more than 1.5 µs of simulation of the protein dimer. The simulations reveal a transition of hSERT from an outward-facing occluded conformation to an inward-facing conformation in a one-substrate-bound state. Simulations with a second substrate in the proposed symport effector site did not lead to conformational changes associated with translocation. The central substrate binding site becomes fully exposed to the cytoplasm leaving both the Na+-ion in the Na2-site and the substrate in direct contact with the cytoplasm through water interactions. The simulations reveal how sodium is released and show indications of early events of substrate transport. The notion that ion dissociation from the Na2-site drives translocation is supported by experimental studies of a Na2-site mutant. Transmembrane helices (TMs) 1 and 6 are identified as the helices involved in the largest movements during transport. PMID:22046120

Strong magnon-phonon coupling in NaFeAs studied by neutron scattering

NASA Astrophysics Data System (ADS)

Li, Yu; Yamani, Zahra; Song, Yu; Zhang, Chenglin; Dai, Pengcheng

We carried on inelastic neutron scattering experiment on the triple axis spectrometer in CNBC in Chalk River. We measured both the phonon and magnon in NaFeAs single crystals and their temperature dependence. Since structural transition temperature (TS) and the magnetic transition temperature (T N) are well separated in NaFeAs, it provides us an unique chance to exclude the consequence or magnetic order and focus on the so called nematic phase. As the previous paper on BaFe2As2, we observed the strong phonon softening nearby the structural transition temperature at very small q (q<0.1). This makes the phonon in NaFeAs deviate from the classical linear dispersion relationship for acoustic phonons. Besides the phonon softening, we also observe phonon hardening at a larger q range when the temperature goes down. This is accompanied by the stiffening of the magnons which can be represented by the linewidth of the low energy magnetic peaks. Our results suggest that there is strong coupling between the phonons and magnons in NaFeAs.

Ion Transport Function of SLC4A11 in Corneal Endothelium

PubMed Central

Jalimarada, Supriya S.; Ogando, Diego G.; Vithana, Eranga N.; Bonanno, Joseph A.

2013-01-01

Purpose. Mutations in SLC4A11, a member of the SLC4 superfamily of bicarbonate transporters, give rise to corneal endothelial cell dystrophies. SLC4A11 is a putative Na+ borate and Na+:OH− transporter. Therefore we ask whether SLC4A11 in corneal endothelium transports borate (B[OH]4−), bicarbonate (HCO3−), or hydroxyl (OH−) anions coupled to Na+. Methods. SLC4A11 expression in cultured primary bovine corneal endothelial cells (BCECs) was determined by semiquantitative PCR, SDS-PAGE/Western blotting, and immunofluorescence staining. Ion transport function was examined by measuring intracellular pH (pHi) or Na+ ([Na+]i) in response to Ringer solutions with/without B(OH)4− or HCO3− after overexpressing or small interfering RNA (siRNA) silencing of SLC4A11. Results. SLC4A11 is localized to the basolateral membrane in BCEC. B(OH)4− (2.5–10 mM) in bicarbonate-free Ringer induced a rapid small acidification (0.01 pH unit) followed by alkalinization (0.05–0.1 pH unit), consistent with diffusion of boric acid into the cell followed by B(OH)4−. However, the rate of B(OH)4−-induced pHi change was unaffected by overexpression of SLC4A11. B(OH)4− did not induce significant changes in resting [Na+i] or the amplitude and rate of acidification caused by Na+ removal. siRNA-mediated knockdown of SLC4A11 (∼70%) did not alter pHi responses to CO2/HCO3−-rich Ringer, Na+-free induced acidification, or the rate of Na+ influx in the presence of bicarbonate. However, in the absence of bicarbonate, siSLC4A11 knockdown significantly decreased the rate (43%) and amplitude (48%) of acidification due to Na+ removal and recovery (53%) upon add-back. Additionally, the rate of acid recovery following NH4+ prepulse was decreased significantly (27%) by SLC4A11 silencing. Conclusions. In corneal endothelium, SLC4A11 displays robust Na+-coupled OH− transport, but does not transport B(OH)4− or HCO3−. PMID:23745003

Na+/H+ exchange activity in the plasma membrane of Arabidopsis.

PubMed

Qiu, Quan-Sheng; Barkla, Bronwyn J; Vera-Estrella, Rosario; Zhu, Jian-Kang; Schumaker, Karen S

2003-06-01

In plants, Na+/H+ exchangers in the plasma membrane are critical for growth in high levels of salt, removing toxic Na+ from the cytoplasm by transport out of the cell. The molecular identity of a plasma membrane Na+/H+ exchanger in Arabidopsis (SOS1) has recently been determined. In this study, immunological analysis provided evidence that SOS1 localizes to the plasma membrane of leaves and roots. To characterize the transport activity of this protein, purified plasma membrane vesicles were isolated from leaves of Arabidopsis. Na+/H+ exchange activity, monitored as the ability of Na to dissipate an established pH gradient, was absent in plants grown without salt. However, exchange activity was induced when plants were grown in 250 mm NaCl and increased with prolonged salt exposure up to 8 d. H+-coupled exchange was specific for Na, because chloride salts of other monovalent cations did not dissipate the pH gradient. Na+/H+ exchange activity was dependent on Na (substrate) concentration, and kinetic analysis indicated that the affinity (apparent Km) of the transporter for Na+ is 22.8 mm. Data from two experimental approaches supports electroneutral exchange (one Na+ exchanged for one proton): (a) no change in membrane potential was measured during the exchange reaction, and (b) Na+/H+ exchange was unaffected by the presence or absence of a membrane potential. Results from this research provide a framework for future studies into the regulation of the plant plasma membrane Na+/H+ exchanger and its relative contribution to the maintenance of cellular Na+ homeostasis during plant growth in salt.

Identification of residues that control Li+ versus Na+ dependent Ca2+ exchange at the transport site of the mitochondrial NCLX.

PubMed

Roy, Soumitra; Dey, Kuntal; Hershfinkel, Michal; Ohana, Ehud; Sekler, Israel

2017-06-01

The Na + /Ca 2+ /Li + exchanger (NCLX) is a member of the Na + /Ca 2+ exchanger family. NCLX is unique in its capacity to transport both Na + and Li + , unlike other members, which are Na + selective. The major aim of this study was twofold, i.e., to identify NCLX residues that confer Li + or Na + selective Ca 2+ transport and map their putative location on NCLX cation transport site. We combined molecular modeling to map transport site of NCLX with euryarchaeal H + /Ca 2+ exchanger, CAX_Af, and fluorescence analysis to monitor Li + versus Na + dependent mitochondrial Ca 2+ efflux of transport site mutants of NCLX in permeabilized cells. Mutation of Asn149, Pro152, Asp153, Gly176, Asn467, Ser468, Gly494 and Asn498 partially or strongly abolished mitochondrial Ca 2+ exchange activity in intact cells. In permeabilized cells, N149A, P152A, D153A, N467Q, S468T and G494S demonstrated normal Li + /Ca 2+ exchange activity but a reduced Na + /Ca 2+ exchange activity. On the other hand, D471A showed dramatically reduced Li + /Ca 2+ exchange, but Na + /Ca 2+ exchange activity was unaffected. Finally, simultaneous mutation of four putative Ca 2+ binding residues was required to completely abolish both Na + /Ca 2+ and Li + /Ca 2+ exchange activities. We identified distinct Na + and Li + selective residues in the NCLX transport site. We propose that functional segregation in Li + and Na + sites reflects the functional properties of NCLX required for Ca 2+ exchange under the unique membrane potential and ion gradient across the inner mitochondrial membrane. The results of this study provide functional insights into the unique Li + and Na + selectivity of the mitochondrial exchanger. This article is part of a Special Issue entitled: ECS Meeting edited by Claus Heizmann, Joachim Krebs and Jacques Haiech. Copyright © 2017 Elsevier B.V. All rights reserved.

Salt stress in Thellungiella halophila activates Na+ transport mechanisms required for salinity tolerance.

PubMed

Vera-Estrella, Rosario; Barkla, Bronwyn J; García-Ramírez, Liliana; Pantoja, Omar

2005-11-01

Salinity is considered one of the major limiting factors for plant growth and agricultural productivity. We are using salt cress (Thellungiella halophila) to identify biochemical mechanisms that enable plants to grow in saline conditions. Under salt stress, the major site of Na+ accumulation occurred in old leaves, followed by young leaves and taproots, with the least accumulation occurring in lateral roots. Salt treatment increased both the H+ transport and hydrolytic activity of salt cress tonoplast (TP) and plasma membrane (PM) H(+)-ATPases from leaves and roots. TP Na(+)/H+ exchange was greatly stimulated by growth of the plants in NaCl, both in leaves and roots. Expression of the PM H(+)-ATPase isoform AHA3, the Na+ transporter HKT1, and the Na(+)/H+ exchanger SOS1 were examined in PMs isolated from control and salt-treated salt cress roots and leaves. An increased expression of SOS1, but no changes in levels of AHA3 and HKT1, was observed. NHX1 was only detected in PM fractions of roots, and a salt-induced increase in protein expression was observed. Analysis of the levels of expression of vacuolar H(+)-translocating ATPase subunits showed no major changes in protein expression of subunits VHA-A or VHA-B with salt treatment; however, VHA-E showed an increased expression in leaf tissue, but not in roots, when the plants were treated with NaCl. Salt cress plants were able to distribute and store Na+ by a very strict control of ion movement across both the TP and PM.

Seed filling in domesticated maize and rice depends on SWEET-mediated hexose transport

USDA-ARS?s Scientific Manuscript database

Carbohydrate import into seeds directly determines seed size and must have been increased through domestication. However, evidence for domestication of sugar translocation and the identity of seed filling transporters remained elusive. Maize ZmSWEET4c, as opposed to its sucrose-transporting homologs...

Transport spectroscopy of coupled donors in silicon nano-transistors

PubMed Central

Moraru, Daniel; Samanta, Arup; Anh, Le The; Mizuno, Takeshi; Mizuta, Hiroshi; Tabe, Michiharu

2014-01-01

The impact of dopant atoms in transistor functionality has significantly changed over the past few decades. In downscaled transistors, discrete dopants with uncontrolled positions and number induce fluctuations in device operation. On the other hand, by gaining access to tunneling through individual dopants, a new type of devices is developed: dopant-atom-based transistors. So far, most studies report transport through dopants randomly located in the channel. However, for practical applications, it is critical to control the location of the donors with simple techniques. Here, we fabricate silicon transistors with selectively nanoscale-doped channels using nano-lithography and thermal-diffusion doping processes. Coupled phosphorus donors form a quantum dot with the ground state split into a number of levels practically equal to the number of coupled donors, when the number of donors is small. Tunneling-transport spectroscopy reveals fine features which can be correlated with the different numbers of donors inside the quantum dot, as also suggested by first-principles simulation results. PMID:25164032

Role of direct electron-phonon coupling across metal-semiconductor interfaces in thermal transport via molecular dynamics.

PubMed

Lin, Keng-Hua; Strachan, Alejandro

2015-07-21

Motivated by significant interest in metal-semiconductor and metal-insulator interfaces and superlattices for energy conversion applications, we developed a molecular dynamics-based model that captures the thermal transport role of conduction electrons in metals and heat transport across these types of interface. Key features of our model, denoted eleDID (electronic version of dynamics with implicit degrees of freedom), are the natural description of interfaces and free surfaces and the ability to control the spatial extent of electron-phonon (e-ph) coupling. Non-local e-ph coupling enables the energy of conduction electrons to be transferred directly to the semiconductor/insulator phonons (as opposed to having to first couple to the phonons in the metal). We characterize the effect of the spatial e-ph coupling range on interface resistance by simulating heat transport through a metal-semiconductor interface to mimic the conditions of ultrafast laser heating experiments. Direct energy transfer from the conduction electrons to the semiconductor phonons not only decreases interfacial resistance but also increases the ballistic transport behavior in the semiconductor layer. These results provide new insight for experiments designed to characterize e-ph coupling and thermal transport at the metal-semiconductor/insulator interfaces.

Methods for stable recording of short-circuit current in a Na+-transporting epithelium.

PubMed

Gondzik, Veronika; Awayda, Mouhamed S

2011-07-01

Epithelial Na(+) transport as measured by a variety of techniques, including the short-circuit current technique, has been described to exhibit a "rundown" phenomenon. This phenomenon manifests as time-dependent decrease of current and resistance and precludes the ability to carry out prolonged experiments aimed at examining the regulation of this transport. We developed methods for prolonged stable recordings of epithelial Na(+) transport using modifications of the short-circuit current technique and commercial Ussing-type chambers. We utilize the polarized MDCK cell line expressing the epithelial Na(+) channel (ENaC) to describe these methods. Briefly, existing commercial chambers were modified to allow continuous flow of Ringer solution and precise control of such flow. Chamber manifolds and associated plumbing were modified to allow precise temperature clamp preventing temperature oscillations. Recording electrodes were modified to eliminate the use of KCl and prevent membrane depolarization from KCl leakage. Solutions utilized standard bicarbonate-based buffers, but all gasses were prehydrated to clamp buffer osmolarity. We demonstrate that these modifications result in measurements of current and resistance that are stable for at least 2 h. We further demonstrate that drifts in osmolarity similar to those obtained before prior to our modifications can lead to a decrease of current and resistance similar to those attributed to rundown.

Vibronic coupling effect on the electron transport through molecules

NASA Astrophysics Data System (ADS)

Tsukada, Masaru; Mitsutake, Kunihiro

2007-03-01

Electron transport through molecular bridges or molecular layers connected to nano-electrodes is determined by the combination of coherent and dissipative processes, controlled by the electron-vibron coupling, transfer integrals between the molecular orbitals, applied electric field and temperature. We propose a novel theoretical approach, which combines ab initio molecular orbital method with analytical many-boson model. As a case study, the long chain model of the thiophene oligomer is solved by a variation approach. Mixed states of moderately extended molecular orbital states mediated and localised by dress of vibron cloud are found as eigen-states. All the excited states accompanied by multiple quanta of vibration can be solved, and the overall carrier transport properties including the conductance, mobility, dissipation spectra are analyzed by solving the master equation with the transition rates estimated by the golden rule. We clarify obtained in a uniform systematic way, how the transport mode changes from a dominantly coherent transport to the dissipative hopping transport.

Characterisation and cloning of a Na(+)-dependent broad-specificity neutral amino acid transporter from NBL-1 cells: a novel member of the ASC/B(0) transporter family.

PubMed

Pollard, Matthew; Meredith, David; McGivan, John D

2002-04-12

Na(+)-dependent neutral amino acid transport into the bovine renal epithelial cell line NBL-1 is catalysed by a broad-specificity transporter originally termed System B(0). This transporter is shown to differ in specificity from the B(0) transporter cloned from JAR cells [J. Biol. Chem. 271 (1996) 18657] in that it interacts much more strongly with phenylalanine. Using probes designed to conserved transmembrane regions of the ASC/B(0) transporter family we have isolated a cDNA encoding the NBL-1 cell System B(0) transporter. When expressed in Xenopus oocytes the clone catalysed Na(+)-dependent alanine uptake which was inhibited by glutamine, leucine and phenylalanine. However, the clone did not catalyse Na(+)-dependent phenylalanine transport, again as in NBL-1 cells. The clone encoded a protein of 539 amino acids; the predicted transmembrane domains were almost identical in sequence to those of the other members of the B(0)/ASC transporter family. Comparison of the sequences of NBL-1 and JAR cell transporters showed some differences near the N-terminus, C-terminus and in the loop between helices 3 and 4. The NBL-1 B(0) transporter is not the same as the renal brush border membrane transporter since it does not transport phenylalanine. Differences in specificity in this protein family arise from relatively small differences in amino acid sequence.

Automated identification of protein-ligand interaction features using Inductive Logic Programming: a hexose binding case study.

PubMed

A Santos, Jose C; Nassif, Houssam; Page, David; Muggleton, Stephen H; E Sternberg, Michael J

2012-07-11

There is a need for automated methods to learn general features of the interactions of a ligand class with its diverse set of protein receptors. An appropriate machine learning approach is Inductive Logic Programming (ILP), which automatically generates comprehensible rules in addition to prediction. The development of ILP systems which can learn rules of the complexity required for studies on protein structure remains a challenge. In this work we use a new ILP system, ProGolem, and demonstrate its performance on learning features of hexose-protein interactions. The rules induced by ProGolem detect interactions mediated by aromatics and by planar-polar residues, in addition to less common features such as the aromatic sandwich. The rules also reveal a previously unreported dependency for residues cys and leu. They also specify interactions involving aromatic and hydrogen bonding residues. This paper shows that Inductive Logic Programming implemented in ProGolem can derive rules giving structural features of protein/ligand interactions. Several of these rules are consistent with descriptions in the literature. In addition to confirming literature results, ProGolem's model has a 10-fold cross-validated predictive accuracy that is superior, at the 95% confidence level, to another ILP system previously used to study protein/hexose interactions and is comparable with state-of-the-art statistical learners.

Insulin-stimulated Na/sup +/ transport in a model renal epithelium: protein synthesis dependence and receptor interactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Blazer-Yost, B.L.; Cox, M.

1987-05-01

The urinary bladder of the toad, Bufo marinus, is a well characterized model of the mammalian distal nephron. Porcine insulin (approx. 0.5-5.0 ..mu..M) stimulates net mucosal to serosal Na/sup +/ flux within 10 minutes of hormone addition. The response is maintained for at least 5 hr and is completely abolished by low doses (10..mu..M) of the epithelial Na/sup +/ channel blocker amiloride. Insulin-stimulated Na/sup +/ transport does not require new protein synthesis since it is actinomycin-D (10..mu..g/ml) insensitive. Also in 3 separate experiments in which epithelial cell proteins were examined by /sup 35/S-methionine labeling, 2-dimensional polyacrylamide gel electrophoresis/autoradiography, no insulinmore » induced proteins were observed. Equimolar concentrations of purified porcine proinsulin and insulin (0.64..mu..M) stimulate Na/sup +/ transport to the same extent. Thus, the putative toad insulin receptor may have different affinity characteristics than those demonstrated for insulin and proinsulin in mammalian tissues. Alternatively, the natriferic action of insulin in toad urinary bladders may be mediated by occupancy of another receptor. Preliminary experiments indicating that nanomolar concentrations of IGF/sub 1/ stimulate Na/sup +/ transport in this tissue support the latter contention.« less

Stochastic analysis of transverse dispersion in density‐coupled transport in aquifers

USGS Publications Warehouse

Welty, Claire; Kane, Allen C.; Kauffman, Leon J.

2003-01-01

Spectral perturbation techniques have been used previously to derive integral expressions for dispersive mixing in concentration‐dependent transport in three‐dimensional, heterogeneous porous media, where fluid density and viscosity are functions of solute concentration. Whereas earlier work focused on evaluating longitudinal dispersivity in isotropic media and incorporating the result in a mean one‐dimensional transport model, the emphasis of this paper is on evaluation of the complete dispersion tensor, including the more general case of anisotropic media. Approximate analytic expressions for all components of the macroscopic dispersivity tensor are derived, and the tensor is shown to be asymmetric. The tensor is separated into its symmetric and antisymmetric parts, where the symmetric part is used to calculate the principal components and principal directions of dispersivity, and the antisymmetric part of the tensor is shown to modify the velocity of the solute body compared to that of the background fluid. An example set of numerical simulations incorporating the tensor illustrates the effect of density‐coupled dispersivity on a sinking plume in an aquifer. The simulations show that the effective transverse vertical spreading in a sinking plume to be significantly greater than would be predicted by a standard density‐coupled transport model that does not incorporate the coupling in the dispersivity tensor.

Nonradiative transport of atomic excitation in Na vapor

NASA Astrophysics Data System (ADS)

Zajonc, Arthur G.; Phelps, A. V.

1981-05-01

Measurements are reported which show the effect of nonradiative losses at a gas-window interface on the backscattered fluorescence intensity for Na vapor at frequencies in the vicinity of the resonance lines near 589 nm. The Na 3P12,32 states are excited with a low-intensity single-mode tunable dye laser at high Na densities and the frequency integral of the backscattered fluorescence intensity in the D1 and D2 lines is measured. As the laser is tuned through resonance, the loss of atomic excitation to the window appears as a sharp decrease in the frequency-integrated fluorescence intensity. For example, at 7×1020 atoms m-3 the fluorescence intensity decreases by a factor of 4 in a frequency interval of 4 GHz. Measured absolute fluorescence intensities versus laser frequency are compared with predictions made using the theory of Hummer and Kunasz which includes both radiative and nonradiative transport processes. The agreement between theory and experiment is remarkably good when one considers that the theory contains only one unknown coefficient, i.e., the reflection coefficient for excited atoms at the windows. In our case the excited atoms are assumed to be completely destroyed at the window.

Na+/H+ Exchange Activity in the Plasma Membrane of Arabidopsis1

PubMed Central

Qiu, Quan-Sheng; Barkla, Bronwyn J.; Vera-Estrella, Rosario; Zhu, Jian-Kang; Schumaker, Karen S.

2003-01-01

In plants, Na+/H+ exchangers in the plasma membrane are critical for growth in high levels of salt, removing toxic Na+ from the cytoplasm by transport out of the cell. The molecular identity of a plasma membrane Na+/H+ exchanger in Arabidopsis (SOS1) has recently been determined. In this study, immunological analysis provided evidence that SOS1 localizes to the plasma membrane of leaves and roots. To characterize the transport activity of this protein, purified plasma membrane vesicles were isolated from leaves of Arabidopsis. Na+/H+ exchange activity, monitored as the ability of Na to dissipate an established pH gradient, was absent in plants grown without salt. However, exchange activity was induced when plants were grown in 250 mm NaCl and increased with prolonged salt exposure up to 8 d. H+-coupled exchange was specific for Na, because chloride salts of other monovalent cations did not dissipate the pH gradient. Na+/H+ exchange activity was dependent on Na (substrate) concentration, and kinetic analysis indicated that the affinity (apparent Km) of the transporter for Na+ is 22.8 mm. Data from two experimental approaches supports electroneutral exchange (one Na+ exchanged for one proton): (a) no change in membrane potential was measured during the exchange reaction, and (b) Na+/H+ exchange was unaffected by the presence or absence of a membrane potential. Results from this research provide a framework for future studies into the regulation of the plant plasma membrane Na+/H+ exchanger and its relative contribution to the maintenance of cellular Na+ homeostasis during plant growth in salt. PMID:12805632

Correlation between uncoupled ATP hydrolysis and heat production by the sarcoplasmic reticulum Ca2+-ATPase: coupling effect of fluoride.

PubMed

Reis, M; Farage, M; de Souza, A C; de Meis, L

2001-11-16

The sarcoplasmic reticulum Ca(2+)-ATPase transports Ca(2+) using the chemical energy derived from ATP hydrolysis. Part of the chemical energy is used to translocate Ca(2+) through the membrane (work) and part is dissipated as heat. The amount of heat produced during catalysis increases after formation of the Ca(2+) gradient across the vesicle membrane. In the absence of gradient (leaky vesicles) the amount of heat produced/mol of ATP cleaved is half of that measured in the presence of the gradient. After formation of the gradient, part of the ATPase activity is not coupled to Ca(2+) transport. We now show that NaF can impair the uncoupled ATPase activity with discrete effect on the ATPase activity coupled to Ca(2+) transport. For the control vesicles not treated with NaF, after formation of the gradient only 20% of the ATP cleaved is coupled to Ca(2+) transport, and the caloric yield of the total ATPase activity (coupled plus uncoupled) is 22.8 kcal released/mol of ATP cleaved. In contrast, the vesicles treated with NaF consume only the ATP needed to maintain the gradient, and the caloric yield of ATP hydrolysis is 3.1 kcal/mol of ATP. The slow ATPase activity measured in vesicles treated with NaF has the same Ca(2+) dependence as the control vesicles. This demonstrates unambiguously that the uncoupled activity is an actual pathway of the Ca(2+)-ATPase rather than a contaminating phosphatase. We conclude that when ATP hydrolysis occurs without coupled biological work most of the chemical energy is dissipated as heat. Thus, uncoupled ATPase activity appears to be the mechanistic feature underlying the ability of the Ca(2+)-ATPase to modulated heat production.

Hierarchical quantum master equation approach to electronic-vibrational coupling in nonequilibrium transport through nanosystems

NASA Astrophysics Data System (ADS)

Schinabeck, C.; Erpenbeck, A.; Härtle, R.; Thoss, M.

2016-11-01

Within the hierarchical quantum master equation (HQME) framework, an approach is presented, which allows a numerically exact description of nonequilibrium charge transport in nanosystems with strong electronic-vibrational coupling. The method is applied to a generic model of vibrationally coupled transport considering a broad spectrum of parameters ranging from the nonadiabatic to the adiabatic regime and including both resonant and off-resonant transport. We show that nonequilibrium effects are important in all these regimes. In particular, in the off-resonant transport regime, the inelastic cotunneling signal is analyzed for a vibrational mode in full nonequilibrium, revealing a complex interplay of different transport processes and deviations from the commonly used G0/2 rule of thumb. In addition, the HQME approach is used to benchmark approximate master equation and nonequilibrium Green's function methods.

Waveguide transport mediated by strong coupling with atoms

NASA Astrophysics Data System (ADS)

Cheng, Mu-Tian; Xu, Jingping; Agarwal, Girish S.

2017-05-01

We investigate single-photon scattering properties in a one-dimensional waveguide coupled to a quantum emitter's chain with dipole-dipole interaction (DDI). The photon transport is extremely sensitive to the location of the evanescently coupled atoms. The analytical expressions of reflection and transmission amplitudes for the chain containing two emitters with DDI are deduced by using a real-space Hamiltonian. Two cases, where the two emitters symmetrically or asymmetrically couple to the waveguide, are discussed in detail. It shows that the reflection and transmission typical spectra split into two peaks due to the DDI. The Fano minimum in the spectra can be used to estimate the strength of the DDI. Furthermore, the DDI makes spectra strongly asymmetric and creates a transmission window in the region where there was zero transmission. The scattering spectra for the chain consisting of multiple emitters are also given. Our key finding is that DDI can broaden the frequency bandwidth for high reflection when the chain consists of many emitters.

Sugar and Glycerol Transport in Saccharomyces cerevisiae.

PubMed

Bisson, Linda F; Fan, Qingwen; Walker, Gordon A

2016-01-01

In Saccharomyces cerevisiae the process of transport of sugar substrates into the cell comprises a complex network of transporters and interacting regulatory mechanisms. Members of the large family of hexose (HXT) transporters display uptake efficiencies consistent with their environmental expression and play physiological roles in addition to feeding the glycolytic pathway. Multiple glucose-inducing and glucose-independent mechanisms serve to regulate expression of the sugar transporters in yeast assuring that expression levels and transporter activity are coordinated with cellular metabolism and energy needs. The expression of sugar transport activity is modulated by other nutritional and environmental factors that may override glucose-generated signals. Transporter expression and activity is regulated transcriptionally, post-transcriptionally and post-translationally. Recent studies have expanded upon this suite of regulatory mechanisms to include transcriptional expression fine tuning mediated by antisense RNA and prion-based regulation of transcription. Much remains to be learned about cell biology from the continued analysis of this dynamic process of substrate acquisition.

Influence of transport energization on the growth yield of Escherichia coli.

PubMed

Muir, M; Williams, L; Ferenci, T

1985-09-01

The growth yields of Escherichia coli on glucose, lactose, galactose, maltose, maltotriose, and maltohexaose were estimated under anaerobic conditions in the absence of electron acceptors. The yields on these substrates exhibited significant differences when measured in carbon-limited chemostats at similar growth rates and compared in terms of grams (dry weight) of cells produced per mole of hexose utilized. Maltohexaose was the most efficiently utilized substrate, and galactose was the least efficiently utilized under these conditions. All these sugars were known to be metabolized to glucose 6-phosphate and produced the same pattern of fermentation products. The differences in growth yields were ascribed to differences in energy costs for transport and phosphorylation of these sugars. A formalized treatment of these factors in determining growth yields was established and used to obtain values for the cost of transport and hence the energy-coupling stoichiometries for the transport of substrates via proton symport and binding-protein-dependent mechanisms in vivo. By this approach, the proton-lactose stoichiometry was found to be 1.1 to 1.8 H+ per lactose, equivalent to approximately 0.5 ATP used per lactose transported. The cost of transporting maltose via a binding-protein-dependent mechanism was considerably higher, being over 1 to 1.2 ATP per maltose or maltodextrin transported. The formalized treatment also permitted estimation of the net ATP yield from the metabolism of these sugars; it was calculated that the growth yield data were consistent with the production of 2.8 to 3.2 ATP in the metabolism of glucose 6-phosphate to fermentation products.

Influence of transport energization on the growth yield of Escherichia coli.

PubMed Central

Muir, M; Williams, L; Ferenci, T

1985-01-01

The growth yields of Escherichia coli on glucose, lactose, galactose, maltose, maltotriose, and maltohexaose were estimated under anaerobic conditions in the absence of electron acceptors. The yields on these substrates exhibited significant differences when measured in carbon-limited chemostats at similar growth rates and compared in terms of grams (dry weight) of cells produced per mole of hexose utilized. Maltohexaose was the most efficiently utilized substrate, and galactose was the least efficiently utilized under these conditions. All these sugars were known to be metabolized to glucose 6-phosphate and produced the same pattern of fermentation products. The differences in growth yields were ascribed to differences in energy costs for transport and phosphorylation of these sugars. A formalized treatment of these factors in determining growth yields was established and used to obtain values for the cost of transport and hence the energy-coupling stoichiometries for the transport of substrates via proton symport and binding-protein-dependent mechanisms in vivo. By this approach, the proton-lactose stoichiometry was found to be 1.1 to 1.8 H+ per lactose, equivalent to approximately 0.5 ATP used per lactose transported. The cost of transporting maltose via a binding-protein-dependent mechanism was considerably higher, being over 1 to 1.2 ATP per maltose or maltodextrin transported. The formalized treatment also permitted estimation of the net ATP yield from the metabolism of these sugars; it was calculated that the growth yield data were consistent with the production of 2.8 to 3.2 ATP in the metabolism of glucose 6-phosphate to fermentation products. PMID:3928598

Influence of cellular and paracellular conductance patterns on epithelial transport and metabolism.

PubMed Central

Essig, A

1982-01-01

Theoretical analysis of transepithelial active Na transport is often based on equivalent electrical circuits comprising discrete parallel active and passive pathways. Recent findings show, however, that Na+ pumps are distributed over the entire basal lateral surface of epithelial cells. This suggests that Na+ that has been actively transported into paracellular channels may to some extent return to the apical (mucosal) bathing solution, depending on the relative conductances of the pathways via the tight junctions and the lateral intercellular spaces. Such circulation, as well as the relative conductance of cellular and paracellular pathways, may have an important influence on the relationships between parameters of transcellular and transepithelial active transport and metabolism. These relationships were examined by equivalent circuit analysis of active Na transport, Na conductance, the electromotive force of Na transport, the "stoichiometry" of transport, and the degree of coupling of transport to metabolism. Although the model is too crude to permit precise quantification, important qualitative differences are predicted between "loose" and "tight" epithelia in the absence and presence of circulation. In contrast, there is no effect on the free energy of metabolic reaction estimated from a linear thermodynamic formalism. Also of interest are implications concerning the experimental evaluation of passive paracellular conductance following abolition of active transport, and the use of the cellular voltage-divider ratio to estimate the relative conductances of apical and basal lateral plasma membranes. PMID:6284264

Hexose Transport in Growing Petunia Pollen Tubes and Characterization of a Pollen-Specific, Putative Monosaccharide Transporter1

PubMed Central

Ylstra, Bauke; Garrido, Dolores; Busscher, Jacqueline; van Tunen, Arjen J.

1998-01-01

We investigated the molecular and physiological processes of sugar uptake and metabolism during pollen tube growth and plant fertilization. In vitro germination assays showed that petunia (Petunia hybrida) pollen can germinate and grow not only in medium containing sucrose (Suc) as a carbon source, but also in medium containing the monosaccharides glucose (Glc) or fructose (Fru). Furthermore, high-performance liquid chromatography analysis demonstrated a rapid and complete conversion of Suc into equimolar amounts of Glc and Fru when pollen was cultured in a medium containing 2% Suc. This indicates the presence of wall-bound invertase activity and uptake of sugars in the form of monosaccharides by the growing pollen tube. A cDNA designated pmt1 (petunia monosaccharide transporter 1), which is highly homologous to plant monosaccharide transporters, was isolated from petunia. Pmt1 belongs to a small gene family and is expressed specifically in the male gametophyte, but not in any other vegetative or floral tissues. Pmt1 is activated after the first pollen mitosis, and high levels of mRNA accumulate in mature and germinating pollen. A model describing the transport of sugars to the style, the conversion of Suc into Glc and Fru, and the active uptake by a monosaccharide transporter into the pollen tube is presented. PMID:9733549

Physiological adjustment to salt stress in Jatropha curcas is associated with accumulation of salt ions, transport and selectivity of K+, osmotic adjustment and K+/Na+ homeostasis.

PubMed

Silva, E N; Silveira, J A G; Rodrigues, C R F; Viégas, R A

2015-09-01

This study assessed the capacity of Jatropha curcas to physiologically adjust to salinity. Seedlings were exposed to increasing NaCl concentrations (25, 50, 75 and 100 mm) for 15 days. Treatment without NaCl was adopted as control. Shoot dry weight was strongly reduced by NaCl, reaching values of 35% to 65% with 25 to 100 mm NaCl. The shoot/root ratio was only affected with 100 mm NaCl. Relative water content (RWC) increased only with 100 mm NaCl, while electrolyte leakage (EL) was much enhanced with 50 mm NaCl. The Na(+) transport rate to the shoot was more affected with 50 and 100 mm NaCl. In parallel, Cl(-) transport rate increased with 75 and 100 mm NaCl, while K(+) transport rate fell from 50 mm to 100 mm NaCl. In roots, Na(+) and Cl(-) transport rates fell slightly only in 50 mm (to Na(+)) and 50 and 100 mm (to Cl(-)) NaCl, while K(+) transport rate fell significantly with increasing NaCl. In general, our data demonstrate that J. curcas seedlings present changes in key physiological processes that allow this species to adjust to salinity. These responses are related to accumulation of Na(+) and Cl(-) in leaves and roots, K(+)/Na(+) homeostasis, transport of K(+) and selectivity (K-Na) in roots, and accumulation of organic solutes contributing to osmotic adjustment of the species. © 2015 German Botanical Society and The Royal Botanical Society of the Netherlands.

Ion sampling and transport in Inductively Coupled Plasma Mass Spectrometry

NASA Astrophysics Data System (ADS)

Farnsworth, Paul B.; Spencer, Ross L.

2017-08-01

Quantitative accuracy and high sensitivity in inductively coupled plasma mass spectrometry (ICP-MS) depend on consistent and efficient extraction and transport of analyte ions from an inductively coupled plasma to a mass analyzer, where they are sorted and detected. In this review we examine the fundamental physical processes that control ion sampling and transport in ICP-MS and compare the results of theory and computerized models with experimental efforts to characterize the flow of ions through plasma mass spectrometers' vacuum interfaces. We trace the flow of ions from their generation in the plasma, into the sampling cone, through the supersonic expansion in the first vacuum stage, through the skimmer, and into the ion optics that deliver the ions to the mass analyzer. At each stage we consider idealized behavior and departures from ideal behavior that affect the performance of ICP-MS as an analytical tool.

Salt Stress in Thellungiella halophila Activates Na+ Transport Mechanisms Required for Salinity Tolerance1

PubMed Central

Vera-Estrella, Rosario; Barkla, Bronwyn J.; García-Ramírez, Liliana; Pantoja, Omar

2005-01-01

Salinity is considered one of the major limiting factors for plant growth and agricultural productivity. We are using salt cress (Thellungiella halophila) to identify biochemical mechanisms that enable plants to grow in saline conditions. Under salt stress, the major site of Na+ accumulation occurred in old leaves, followed by young leaves and taproots, with the least accumulation occurring in lateral roots. Salt treatment increased both the H+ transport and hydrolytic activity of salt cress tonoplast (TP) and plasma membrane (PM) H+-ATPases from leaves and roots. TP Na+/H+ exchange was greatly stimulated by growth of the plants in NaCl, both in leaves and roots. Expression of the PM H+-ATPase isoform AHA3, the Na+ transporter HKT1, and the Na+/H+ exchanger SOS1 were examined in PMs isolated from control and salt-treated salt cress roots and leaves. An increased expression of SOS1, but no changes in levels of AHA3 and HKT1, was observed. NHX1 was only detected in PM fractions of roots, and a salt-induced increase in protein expression was observed. Analysis of the levels of expression of vacuolar H+-translocating ATPase subunits showed no major changes in protein expression of subunits VHA-A or VHA-B with salt treatment; however, VHA-E showed an increased expression in leaf tissue, but not in roots, when the plants were treated with NaCl. Salt cress plants were able to distribute and store Na+ by a very strict control of ion movement across both the TP and PM. PMID:16244148

Regulators of Slc4 bicarbonate transporter activity

PubMed Central

Thornell, Ian M.; Bevensee, Mark O.

2015-01-01

The Slc4 family of transporters is comprised of anion exchangers (AE1-4), Na+-coupled bicarbonate transporters (NCBTs) including electrogenic Na/bicarbonate cotransporters (NBCe1 and NBCe2), electroneutral Na/bicarbonate cotransporters (NBCn1 and NBCn2), and the electroneutral Na-driven Cl-bicarbonate exchanger (NDCBE), as well as a borate transporter (BTR1). These transporters regulate intracellular pH (pHi) and contribute to steady-state pHi, but are also involved in other physiological processes including CO2 carriage by red blood cells and solute secretion/reabsorption across epithelia. Acid-base transporters function as either acid extruders or acid loaders, with the Slc4 proteins moving HCO−3 either into or out of cells. According to results from both molecular and functional studies, multiple Slc4 proteins and/or associated splice variants with similar expected effects on pHi are often found in the same tissue or cell. Such apparent redundancy is likely to be physiologically important. In addition to regulating pHi, a HCO−3 transporter contributes to a cell's ability to fine tune the intracellular regulation of the cotransported/exchanged ion(s) (e.g., Na+ or Cl−). In addition, functionally similar transporters or splice variants with different regulatory profiles will optimize pH physiology and solute transport under various conditions or within subcellular domains. Such optimization will depend on activated signaling pathways and transporter expression profiles. In this review, we will summarize and discuss both well-known and more recently identified regulators of the Slc4 proteins. Some of these regulators include traditional second messengers, lipids, binding proteins, autoregulatory domains, and less conventional regulators. The material presented will provide insight into the diversity and physiological significance of multiple members within the Slc4 gene family. PMID:26124722

Regulators of Slc4 bicarbonate transporter activity.

PubMed

Thornell, Ian M; Bevensee, Mark O

2015-01-01

The Slc4 family of transporters is comprised of anion exchangers (AE1-4), Na(+)-coupled bicarbonate transporters (NCBTs) including electrogenic Na/bicarbonate cotransporters (NBCe1 and NBCe2), electroneutral Na/bicarbonate cotransporters (NBCn1 and NBCn2), and the electroneutral Na-driven Cl-bicarbonate exchanger (NDCBE), as well as a borate transporter (BTR1). These transporters regulate intracellular pH (pHi) and contribute to steady-state pHi, but are also involved in other physiological processes including CO2 carriage by red blood cells and solute secretion/reabsorption across epithelia. Acid-base transporters function as either acid extruders or acid loaders, with the Slc4 proteins moving HCO(-) 3 either into or out of cells. According to results from both molecular and functional studies, multiple Slc4 proteins and/or associated splice variants with similar expected effects on pHi are often found in the same tissue or cell. Such apparent redundancy is likely to be physiologically important. In addition to regulating pHi, a HCO(-) 3 transporter contributes to a cell's ability to fine tune the intracellular regulation of the cotransported/exchanged ion(s) (e.g., Na(+) or Cl(-)). In addition, functionally similar transporters or splice variants with different regulatory profiles will optimize pH physiology and solute transport under various conditions or within subcellular domains. Such optimization will depend on activated signaling pathways and transporter expression profiles. In this review, we will summarize and discuss both well-known and more recently identified regulators of the Slc4 proteins. Some of these regulators include traditional second messengers, lipids, binding proteins, autoregulatory domains, and less conventional regulators. The material presented will provide insight into the diversity and physiological significance of multiple members within the Slc4 gene family.

Parameterization and Modeling of Coupled Heat and Mass Transport in the Vadose Zone

NASA Astrophysics Data System (ADS)

Mohanty, B.; Yang, Z.

2016-12-01

The coupled heat and mass transport in the vadose zone is essentially a multiphysics issue. Addressing this issue appropriately has remarkable impacts on soil physical, chemical and biological processes. To data, most coupled heat and water transport modeling has focused on the interactions between liquid water, water vapor and heat transport in homogeneous and layered soils. Comparatively little work has been done on structured soils where preferential infiltration and evaporation flow occurs. Moreover, the traditional coupled heat and water model usually neglects the nonwetting phase air flow, which was found to be significant in the state-of-the-art modeling framework for coupled heat and water transport investigation. However, the parameterizations for the nonwetting phase air permeability largely remain elusive so far. In order to address the above mentioned limitations, this study aims to develop and validate a predictive multiphysics modeling framework for coupled soil heat and water transport in the heterogeneous shallow subsurface. To this end, the following research work is specifically conducted: (a) propose an improved parameterization to better predict the nonwetting phase relative permeability; (b) determine the dynamics, characteristics and processes of simultaneous soil moisture and heat movement in homogeneous and layered soils; and (c) develop a nonisothermal dual permeability model for heterogeneous structured soils. The results of our studies showed that: (a) the proposed modified nonwetting phase relative permeability models are much more accurate, which can be adopted for better parameterization in the subsequent nonisothermal two phase flow models; (b) the isothermal liquid film flow, nonwetting phase gas flow and liquid-vapor phase change non-equilibrium effects are significant in the arid and semiarid environments (Riverside, California and Audubon, Arizona); and (c) the developed nonisothermal dual permeability model is capable of

Intestinal transport: studies with isolated epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kimmich, G.A.

1979-12-01

Isolated intestinal epithelial cells have been extremely useful for characterizing the nature of intestinal absorption processes and for providing insight into the energetics of Na/sup +/-dependent transport systems. This report describes a number of experimental approaches which have been used for investigating the specific epithelial transport systems involved in sugar absorption, but provides information which ultimately should prove useful for characterizing a number of different intestinal transport events. Similar experiments should also prove useful for exploring the effect of environmental agents on the function of intestinal tissue. In the case of sugars, net absorption is accomplished via a mucosal, Na/supmore » +/-dependent concentrative transport system acting in sequence with a passive serosal system which does not require Na/sup +/. The serosal system limits the full gradient-forming capability of the muscosal system. Agents such as phloretin or cytochalasin B which inhibit serosal transport allow the cells to establish sugar gradients as high as 70 fold in contrast to 10 to 15 fold gradients observed for control cells. Sevety-fold sugar gradients cannot be explained in terms of the energy available in the electrochemical potential for Na/sup +/ if the Na/sub 2/: sugar coupling stoichiometry is 1:1 as commonly assumed. New information indicates that the true Na/sup +/:sugar stoichiometry is in fact 2:1. Flow of two Na/sup +/ ions per sugar molecule down the transmembrane electrochemical potential for Na/sup +/ provides more than sufficient energy to account for observed 70 fold sugar gradients. If flow of sugar by other routes could be completely inhibited, theoretical sugar gradients as high as 400 could be achieved assuming that the cells maintain a membrane potential of -36 mV as measured for intact tissue.« less

Coupled Modeling of Flow, Transport, and Deformation during Hydrodynamically Unstable Displacement in Fractured Rocks

NASA Astrophysics Data System (ADS)

Jha, B.; Juanes, R.

2015-12-01

Coupled processes of flow, transport, and deformation are important during production of hydrocarbons from oil and gas reservoirs. Effective design and implementation of enhanced recovery techniques such as miscible gas flooding and hydraulic fracturing requires modeling and simulation of these coupled proceses in geologic porous media. We develop a computational framework to model the coupled processes of flow, transport, and deformation in heterogeneous fractured rock. We show that the hydrocarbon recovery efficiency during unstable displacement of a more viscous oil with a less viscous fluid in a fractured medium depends on the mechanical state of the medium, which evolves due to permeability alteration within and around fractures. We show that fully accounting for the coupling between the physical processes results in estimates of the recovery efficiency in agreement with observations in field and lab experiments.

Transport regimes spanning magnetization-coupling phase space

NASA Astrophysics Data System (ADS)

Baalrud, Scott D.; Daligault, Jérôme

2017-10-01

The manner in which transport properties vary over the entire parameter-space of coupling and magnetization strength is explored. Four regimes are identified based on the relative size of the gyroradius compared to other fundamental length scales: the collision mean free path, Debye length, distance of closest approach, and interparticle spacing. Molecular dynamics simulations of self-diffusion and temperature anisotropy relaxation spanning the parameter space are found to agree well with the predicted boundaries. Comparison with existing theories reveals regimes where they succeed, where they fail, and where no theory has yet been developed.

Coupled-resonator waveguide perfect transport single-photon by interatomic dipole-dipole interaction

NASA Astrophysics Data System (ADS)

Yan, Guo-an; Lu, Hua; Qiao, Hao-xue; Chen, Ai-xi; Wu, Wan-qing

2018-06-01

We theoretically investigate single-photon coherent transport in a one-dimensional coupled-resonator waveguide coupled to two quantum emitters with dipole-dipole interactions. The numerical simulations demonstrate that the transmission spectrum of the photon depends on the two atoms dipole-dipole interactions and the photon-atom couplings. The dipole-dipole interactions may change the dip positions in the spectra and the coupling strength may broaden the frequency band width in the transmission spectrum. We further demonstrate that the typical transmission spectra split into two dips due to the dipole-dipole interactions. This phenomenon may be used to manufacture new quantum waveguide devices.

Lysine 300 is essential for stability but not for electrogenic transport of the Escherichia coli NhaA Na+/H+ antiporter

PubMed Central

Călinescu, Octavian; Dwivedi, Manish; Patiño-Ruiz, Miyer; Padan, Etana; Fendler, Klaus

2017-01-01

Na+/H+ antiporters are located in the cytoplasmic and intracellular membranes and play crucial roles in regulating intracellular pH, Na+, and volume. The NhaA antiporter of Escherichia coli is the best studied member of the Na+/H+ exchanger family and a model system for all related Na+/H+ exchangers, including eukaryotic representatives. Several amino acid residues are important for the transport activity of NhaA, including Lys-300, a residue that has recently been proposed to carry one of the two H+ ions that NhaA exchanges for one Na+ ion during one transport cycle. Here, we sought to characterize the effects of mutating Lys-300 of NhaA to amino acid residues containing side chains of different polarity and length (i.e. Ala, Arg, Cys, His, Glu, and Leu) on transporter stability and function. Salt resistance assays, acridine-orange fluorescence dequenching, solid supported membrane-based electrophysiology, and differential scanning fluorometry were used to characterize Na+ and H+ transport, charge translocation, and thermal stability of the different variants. These studies revealed that NhaA could still perform electrogenic Na+/H+ exchange even in the absence of a protonatable residue at the Lys-300 position. However, all mutants displayed lower thermal stability and reduced ion transport activity compared with the wild-type enzyme, indicating the critical importance of Lys-300 for optimal NhaA structural stability and function. On the basis of these experimental data, we propose a tentative mechanism integrating the functional and structural role of Lys-300. PMID:28330875

Lysine 300 is essential for stability but not for electrogenic transport of the Escherichia coli NhaA Na+/H+ antiporter.

PubMed

Călinescu, Octavian; Dwivedi, Manish; Patiño-Ruiz, Miyer; Padan, Etana; Fendler, Klaus

2017-05-12

Na + /H + antiporters are located in the cytoplasmic and intracellular membranes and play crucial roles in regulating intracellular pH, Na + , and volume. The NhaA antiporter of Escherichia coli is the best studied member of the Na + /H + exchanger family and a model system for all related Na + /H + exchangers, including eukaryotic representatives. Several amino acid residues are important for the transport activity of NhaA, including Lys-300, a residue that has recently been proposed to carry one of the two H + ions that NhaA exchanges for one Na + ion during one transport cycle. Here, we sought to characterize the effects of mutating Lys-300 of NhaA to amino acid residues containing side chains of different polarity and length ( i.e. Ala, Arg, Cys, His, Glu, and Leu) on transporter stability and function. Salt resistance assays, acridine-orange fluorescence dequenching, solid supported membrane-based electrophysiology, and differential scanning fluorometry were used to characterize Na + and H + transport, charge translocation, and thermal stability of the different variants. These studies revealed that NhaA could still perform electrogenic Na + /H + exchange even in the absence of a protonatable residue at the Lys-300 position. However, all mutants displayed lower thermal stability and reduced ion transport activity compared with the wild-type enzyme, indicating the critical importance of Lys-300 for optimal NhaA structural stability and function. On the basis of these experimental data, we propose a tentative mechanism integrating the functional and structural role of Lys-300. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Automated identification of protein-ligand interaction features using Inductive Logic Programming: a hexose binding case study

PubMed Central

2012-01-01

Background There is a need for automated methods to learn general features of the interactions of a ligand class with its diverse set of protein receptors. An appropriate machine learning approach is Inductive Logic Programming (ILP), which automatically generates comprehensible rules in addition to prediction. The development of ILP systems which can learn rules of the complexity required for studies on protein structure remains a challenge. In this work we use a new ILP system, ProGolem, and demonstrate its performance on learning features of hexose-protein interactions. Results The rules induced by ProGolem detect interactions mediated by aromatics and by planar-polar residues, in addition to less common features such as the aromatic sandwich. The rules also reveal a previously unreported dependency for residues cys and leu. They also specify interactions involving aromatic and hydrogen bonding residues. This paper shows that Inductive Logic Programming implemented in ProGolem can derive rules giving structural features of protein/ligand interactions. Several of these rules are consistent with descriptions in the literature. Conclusions In addition to confirming literature results, ProGolem’s model has a 10-fold cross-validated predictive accuracy that is superior, at the 95% confidence level, to another ILP system previously used to study protein/hexose interactions and is comparable with state-of-the-art statistical learners. PMID:22783946

Sarcolemmal localisation of Na+/H+ exchange and Na+–HCO3− co-transport influences the spatial regulation of intracellular pH in rat ventricular myocytes

PubMed Central

Garciarena, Carolina D; Ma, Yu-ling; Swietach, Pawel; Huc, Laurence; Vaughan-Jones, Richard D

2013-01-01

Membrane acid extrusion by Na+/H+ exchange (NHE1) and Na+–HCO3− co-transport (NBC) is essential for maintaining a low cytoplasmic [H+] (∼60 nm, equivalent to an intracellular pH (pHi) of 7.2). This protects myocardial function from the high chemical reactivity of H+ ions, universal end-products of metabolism. We show here that, in rat ventricular myocytes, fluorescent antibodies map the NBC isoforms NBCe1 and NBCn1 to lateral sarcolemma, intercalated discs and transverse tubules (t-tubules), while NHE1 is absent from t-tubules. This unexpected difference matches functional measurements of pHi regulation (using AM-loaded SNARF-1, a pH fluorophore). Thus, myocyte detubulation (by transient exposure to 1.5 m formamide) reduces global acid extrusion on NBC by 40%, without affecting NHE1. Similarly, confocal pHi imaging reveals that NBC stimulation induces spatially uniform pHi recovery from acidosis, whereas NHE1 stimulation induces pHi non-uniformity during recovery (of ∼0.1 units, for 2–3 min), particularly at the ends of the cell where intercalated discs are commonly located, and where NHE1 immunostaining is prominent. Mathematical modelling shows that this induction of local pHi microdomains is favoured by low cytoplasmic H+ mobility and long H+ diffusion distances, particularly to surface NHE1 transporters mediating high membrane flux. Our results provide the first evidence for a spatial localisation of [H+]i regulation in ventricular myocytes, suggesting that, by guarding pHi, NHE1 preferentially protects gap junctional communication at intercalated discs, while NBC locally protects t-tubular excitation–contraction coupling. PMID:23420656

Phosphoglucoisomerase-catalyzed interconversion of hexose phosphates. Study by 13C NMR of proton and deuteron exchange.

PubMed

Malaisse, W J; Liemans, V; Malaisse-Lagae, F; Ottinger, R; Willem, R

1991-05-15

The exchange of protons and deuterons by phosphoglucoisomerase during the single passage conversion of D-[2-13C,1-2H]fructose 6-phosphate in H2O or D-[2-13C]fructose 6-phosphate in D2O to D-[2-13C]glucose 6-phosphate, as coupled with the further generation of 6-phospho-D-[2-13C]gluconate in the presence of excess glucose-6-phosphate dehydrogenase was investigated by 13C NMR spectroscopy of the latter metabolite. In H2O, the intramolecular deuteron transfer from the C1 of D-fructose 6-phosphate to the C2 of D-glucose 6-phosphate amounted to 65%, a value only slightly lower than the 72% intramolecular proton transfer in D2O. Both percentages, especially the latter one, were lower than those previously recorded during the single passage conversion of D-[1-13C,2-2H]glucose 6-phosphate in H2O or D-[1-13C]glucose 6-phosphate in D2O to D-fructose 6-phosphate and then to D-fructose 1,6-bisphosphate. These differences indicate that the sequence of interactions between the hexose esters and the binding sites of phosphoglucoisomerase is not strictly in mirror image during, respectively, the conversion of the aldose phosphate to ketose phosphate and the opposite process.

Mechanisms of dietary Cu uptake in freshwater rainbow trout: evidence for Na-assisted Cu transport and a specific metal carrier in the intestine.

PubMed

Nadella, Sunita Rao; Grosell, Martin; Wood, Chris M

2007-05-01

Copper (Cu) is both a vital nutrient and a potent toxicant. The objective of this study was to analyze the mechanistic nature of intestinal Cu transport in rainbow trout using radiolabeled Cu (64Cu) and an in vitro gut sac technique. Reduction of mucosal NaCl levels inhibited Cu transport while increase caused stimulation; Na(2)SO(4) had an identical effect, implicating Na(+) rather than the anion. These responses were unrelated to solvent drag, osmotic pressure or changes in transepithelial potential. The presence of elevated luminal Ag stimulated Cu and Na(+) uptake. Phenamil caused a partial inhibition of both Cu and Na(+) uptake while hypercapnia stimulated Na(+) and Cu transport. Cu uptake was sensitive to luminal pH and inhibited by a tenfold excess of Fe and Zn. These factors had no effect on Na(+ )uptake. On the basis of these results we propose a novel Na(+)-assisted mechanism of Cu uptake wherein the Na(+) gradient stimulates an increase in the H(+) concentration of the brushborder creating a suitable microenvironment for the effective transport of Cu via either DMT1 or Ctr1.

Na+-independent transporters, LAT-2 and b0,+, exchange L-DOPA with neutral and basic amino acids in two clonal renal cell lines.

PubMed

Gomes, P; Soares-da-Silva, P

2002-03-15

The present study examined the functional characteristics of L-DOPA transporters in two functionally different clonal subpopulations of opossum kidney (OKLC and OKHC) cells. The uptake of L-DOPA was largely Na+-independent, though in OKHC cells a minor component (approximately 15%) required extracellular Na+. At least two Na+-independent transporters appear to be involved in L-DOPA uptake. One of these transporters has a broad specificity for small and large neutral amino acids, is stimulated by acid pH and inhibited by 2-aminobicyclo(2,2,l)-heptane-2-carboxylic acid (BCH; OKLC, Ki = 291 mM; OKHC, Ki = 380 mM). The other Na+-independent transporter binds neutral and basic amino acids and also recognizes the di-amino acid cystine. [14C]-L-DOPA efflux from OKLC and OKHC cells over 12 min corresponded to a small amount of intracellular [14C]-L-DOPA. L-Leucine, nonlabelled L-DOPA, BCH and L-arginine, stimulated the efflux of [14C]-L-DOPA in a Na+-independent manner. It is suggested that L-DOPA uses at least two major transporters, systems LAT-2 and b0,+. The transport of L-DOPA by LAT-2 corresponds to a Na+-independent transporter with a broad specificity for small and large neutral amino acids, stimulated by acid pH and inhibited by BCH. The transport of L-DOPA by system b0,+ is a Na+-independent transporter for neutral and basic amino acids that also recognizes cystine. LAT-2 was found equally important at the apical and basolateral membranes, whereas system b0,+ had a predominant distribution in apical membranes.

Na(+) transport, and the E(1)P-E(2)P conformational transition of the Na(+)/K(+)-ATPase.

PubMed Central

Babes, A; Fendler, K

2000-01-01

We have used admittance analysis together with the black lipid membrane technique to analyze electrogenic reactions within the Na(+) branch of the reaction cycle of the Na(+)/K(+)-ATPase. ATP release by flash photolysis of caged ATP induced changes in the admittance of the compound membrane system that are associated with partial reactions of the Na(+)/K(+)-ATPase. Frequency spectra and the Na(+) dependence of the capacitive signal are consistent with an electrogenic or electroneutral E(1)P <--> E(2)P conformational transition which is rate limiting for a faster electrogenic Na(+) dissociation reaction. We determine the relaxation rate of the rate-limiting reaction and the equilibrium constants for both reactions at pH 6.2-8.5. The relaxation rate has a maximum value at pH 7.4 (approximately 320 s(-1)), which drops to acidic (approximately 190 s(-1)) and basic (approximately 110 s(-1)) pH. The E(1)P <--> E(2)P equilibrium is approximately at a midpoint position at pH 6.2 (equilibrium constant approximately 0.8) but moves more to the E(1)P side at basic pH 8.5 (equilibrium constant approximately 0.4). The Na(+) affinity at the extracellular binding site decreases from approximately 900 mM at pH 6.2 to approximately 200 mM at pH 8.5. The results suggest that during Na(+) transport the free energy supplied by the hydrolysis of ATP is mainly used for the generation of a low-affinity extracellular Na(+) discharge site. Ionic strength and lyotropic anions both decrease the relaxation rate. However, while ionic strength does not change the position of the conformational equilibrium E(1)P <--> E(2)P, lyotropic anions shift it to E(1)P. PMID:11053130

Ascorbate transport in pig coronary artery smooth muscle: Na(+) removal and oxidative stress increase loss of accumulated cellular ascorbate.

PubMed

Holmes, M E; Samson, S E; Wilson, J X; Dixon, S J; Grover, A K

2000-01-01

Pig deendothelialized coronary artery rings and smooth muscle cells cultured from them accumulated ascorbate from medium containing Na(+). The accumulated material was determined to be ascorbate using high-performance liquid chromatography. We further characterized ascorbate uptake in the cultured cells. The data fitted best with a Hill coefficient of 1 for ascorbate (K(asc) = 22 +/- 2 microM) and 2 for Na(+) (K(Na) = 84 +/- 10 mM). The anion transport inhibitors sulfinpyrazone and 4,4'-diisothiocyanatostilbene-2,2'-disulfonate (DIDS) inhibited the uptake. Transferring cultured cells loaded with (14)C-ascorbate into an ascorbate-free solution resulted in a biphasic loss of radioactivity - an initial sulfinpyrazone-insensitive faster phase and a late sulfinpyrazone-sensitive slower phase. Transferring loaded cells into a Na(+)-free medium increased the loss in the initial phase in a sulfinpyrazone-sensitive manner, suggesting that the ascorbate transporter is bidirectional. Including peroxide or superoxide in the solution increased the loss of radioactivity. Thus, ascorbate accumulated in coronary artery smooth muscle cells by a Na(+)-dependent transporter was lost in an ascorbate-free solution, and the loss was increased by removing Na(+) from the medium or by oxidative stress. Copyright 2000 S. Karger AG, Basel

Molecular properties of the SLC13 family of dicarboxylate and sulfate transporters

PubMed Central

Pajor, Ana M.

2006-01-01

The SLC13 gene family consists of five members in humans, with corresponding orthologs from different vertebrate species. All five genes code for sodium-coupled transporters that are found on the plasma membrane. Two of the transporters, NaS1 and NaS2, carry substrates such as sulfate, selenate and thiosulfate. The other members of the family (NaDC1, NaDC3, and NaCT) are transporters for di- and tri-carboxylates including succinate, citrate and α-ketoglutarate. The SLC13 transporters from vertebrates are electrogenic and they produce inward currents in the presence of sodium and substrate. Substrate-independent leak currents have also been described. Structure–function studies have identified the carboxy terminal half of these proteins as the most important for determining function. Transmembrane helices 9 and 10 may form part of the substrate permeation pathway and participate in conformational changes during the transport cycle. This review also discusses new members of the SLC13 superfamily that exhibit both sodium-dependent and sodium-independent transport mechanisms. The Indy protein from Drosophila, involved in determining lifespan, and the plant vacuolar malate transporter are both sodium-independent dicarboxylate transporters, possibly acting as exchangers. The purpose of this review is to provide an update on new advances in this gene family, particularly on structure–function studies and new members of the family. PMID:16211368

Role of NH3 and NH4+ transporters in renal acid-base transport.

PubMed

Weiner, I David; Verlander, Jill W

2011-01-01

Renal ammonia excretion is the predominant component of renal net acid excretion. The majority of ammonia excretion is produced in the kidney and then undergoes regulated transport in a number of renal epithelial segments. Recent findings have substantially altered our understanding of renal ammonia transport. In particular, the classic model of passive, diffusive NH3 movement coupled with NH4+ "trapping" is being replaced by a model in which specific proteins mediate regulated transport of NH3 and NH4+ across plasma membranes. In the proximal tubule, the apical Na+/H+ exchanger, NHE-3, is a major mechanism of preferential NH4+ secretion. In the thick ascending limb of Henle's loop, the apical Na+-K+-2Cl- cotransporter, NKCC2, is a major contributor to ammonia reabsorption and the basolateral Na+/H+ exchanger, NHE-4, appears to be important for basolateral NH4+ exit. The collecting duct is a major site for renal ammonia secretion, involving parallel H+ secretion and NH3 secretion. The Rhesus glycoproteins, Rh B Glycoprotein (Rhbg) and Rh C Glycoprotein (Rhcg), are recently recognized ammonia transporters in the distal tubule and collecting duct. Rhcg is present in both the apical and basolateral plasma membrane, is expressed in parallel with renal ammonia excretion, and mediates a critical role in renal ammonia excretion and collecting duct ammonia transport. Rhbg is expressed specifically in the basolateral plasma membrane, and its role in renal acid-base homeostasis is controversial. In the inner medullary collecting duct (IMCD), basolateral Na+-K+-ATPase enables active basolateral NH4+ uptake. In addition to these proteins, several other proteins also contribute to renal NH3/NH4+ transport. The role and mechanisms of these proteins are discussed in depth in this review.

A Barley Efflux Transporter Operates in a Na+-Dependent Manner, as Revealed by a Multidisciplinary Platform[OPEN

PubMed Central

Nagarajan, Yagnesh; Rongala, Jay; Luang, Sukanya; Shadiac, Nadim; Sutton, Tim; Tyerman, Stephen D.; McPhee, Gordon; Voelcker, Nicolas H.; Lee, Jung-Goo

2016-01-01

Plant growth and survival depend upon the activity of membrane transporters that control the movement and distribution of solutes into, around, and out of plants. Although many plant transporters are known, their intrinsic properties make them difficult to study. In barley (Hordeum vulgare), the root anion-permeable transporter Bot1 plays a key role in tolerance to high soil boron, facilitating the efflux of borate from cells. However, its three-dimensional structure is unavailable and the molecular basis of its permeation function is unknown. Using an integrative platform of computational, biophysical, and biochemical tools as well as molecular biology, electrophysiology, and bioinformatics, we provide insight into the origin of transport function of Bot1. An atomistic model, supported by atomic force microscopy measurements, reveals that the protein folds into 13 transmembrane-spanning and five cytoplasmic α-helices. We predict a trimeric assembly of Bot1 and the presence of a Na+ ion binding site, located in the proximity of a pore that conducts anions. Patch-clamp electrophysiology of Bot1 detects Na+-dependent polyvalent anion transport in a Nernstian manner with channel-like characteristics. Using alanine scanning, molecular dynamics simulations, and transport measurements, we show that conductance by Bot1 is abolished by removal of the Na+ ion binding site. Our data enhance the understanding of the permeation functions of Bot1. PMID:26672067

Normal and anomalous transport phenomena in two-dimensional NaCl, MoS2 and honeycomb surfaces

NASA Astrophysics Data System (ADS)

Mbemmo, A. M. Fopossi; Kenmoé, G. Djuidjé; Kofané, T. C.

2018-04-01

Understanding the effects of anisotropy and substrate shape on the stochastic processes is critically needed for the improvement of the quality of the transport information. The effect of biharmonic force on the transport phenomena of a particle in two-dimensional is investigated in the framework of three representative substrate lattices: NaCl, MoS2 and honeycomb. We focus on the particles drift velocity, to characterize the transport properties in the system. Normal and anomalous transport are identified for a particular set of the system parameters such as the biharmonic parameter, the bias force, the phase-lag of two signals, as well as the noise amplitude. According to the direction ψ where the bias force is applied, we determine the biharmonic parameter ɛ for the presence of anomalous transport and show that for the NaCl surface, the anomalous transport is observed for 2 < ɛ < 10. For the MoS2 surface, it appears at monochromatic driven (ɛ = 0) and for 3 < ɛ < 9. In particular for the honeycomb surface anomalous transport is generated for 0 ⩽ ɛ < 6 only when ψ > 30 °.

γ-Adducin Stimulates the Thiazide-sensitive NaCl Cotransporter

PubMed Central

Dimke, Henrik; San-Cristobal, Pedro; de Graaf, Mark; Lenders, Jacques W.; Deinum, Jaap; Hoenderop, Joost G.J.

2011-01-01

The thiazide-sensitive NaCl cotransporter (NCC) plays a key role in renal salt reabsorption and the determination of systemic BP, but the molecular mechanisms governing the regulation of NCC are not completely understood. Here, through pull-down experiments coupled to mass spectrometry, we found that γ-adducin interacts with the NCC transporter. γ-Adducin colocalized with NCC to the distal convoluted tubule. 22Na+ uptake experiments in the Xenopus laevis oocyte showed that γ-adducin stimulated NCC activity in a dose-dependent manner, an effect that occurred upstream from With No Lysine (WNK) 4 kinase. The binding site of γ-adducin mapped to the N terminus of NCC and encompassed three previously reported phosphorylation sites. Supporting this site of interaction, competition with the N-terminal domain of NCC abolished the stimulatory effect of γ-adducin on the transporter. γ-Adducin failed to increase NCC activity when these phosphorylation sites were constitutively inactive or active. In addition, γ-adducin bound only to the dephosphorylated N terminus of NCC. Taken together, our observations suggest that γ-adducin dynamically regulates NCC, likely by amending the phosphorylation state, and consequently the activity, of the transporter. These data suggest that γ-adducin may influence BP homeostasis by modulating renal NaCl transport. PMID:21164023

[Effect of NaHCO3 stress on uptake and transportation of Na+, K+ and Ca2+ in three shrub species].

PubMed

Mao, Gui-Lian; Li, Guo-Qi; Xu, Xing; Zhang, Xin-Xue

2014-03-01

We detected absorption and transportation of ions in the leaves of Atriplex nummularia, Atriplex canescens and Lycium barbarum under NaHCO3 stress (300 mmol x L(-1)) by using atomic absorption spectrophotometry and non-invasive ion flux measurement. The results showed that leaves of the A. nummularia, A. canescens and L. barbarum exhibited a high capacity to induce the Na+ accumulation when compared with that of control. The higher the concentration of NaHCO3 treatment, the more Na+ accumulated in the leaves of the three plants under experimental condition. L. barbarum showed a higher Na+ efflux in the mesophyll cells, whereas A. nummularia and A. canescens showed a relative lower efflux. A lower K+ content and a higher Na+/K+ ratio were detected in leaves of A. nummularia and L. barbarum. However, a higher K+ content and a lower Na+/K+ ratio were seen in leaves of A. canescens. Due to induction of Ca2+ efflux under the NaHCO3 treatment, a lower Ca2+ content and a higher Na+/Ca2+ ratio were observed in L. barbarum. On the contrary, a higher Ca2+ influx was observed in A. nummularia and A. canescens. These results suggested that the three shrubs species had different Na+ segmentation strategies. The accumulation of Na+ inhibited Ca2+ absorption in leaves of L. barbarum, while in the A. nummularia and A. canescens, Ca2+ influx induced [Ca2+]cyt which preserved a less-depolarized PM and then inhibited K efflux. The maintaining of cellular K+/Na+ homeostasis in A. nummularia and A. canescens might be achieved by the induction of [Ca2+]cyt under the NaHCO3 treatment.

First evidence of epithelial transport in tardigrades: a comparative investigation of organic anion transport.

PubMed

Halberg, Kenneth Agerlin; Møbjerg, Nadja

2012-02-01

We investigated transport of the organic anion Chlorophenol Red (CPR) in the tardigrade Halobiotus crispae using a new method for quantifying non-fluorescent dyes. We compared the results acquired from the tardigrade with CPR transport data obtained from Malpighian tubules of the desert locust Schistocerca gregaria. CPR accumulated in the midgut lumen of H. crispae, indicating that organic anion transport takes place here. Our results show that CPR transport is inhibited by the mitochondrial un-coupler DNP (1 mmol l(-1); 81% reduction), the Na(+)/K(+)-ATPase inhibitor ouabain (10 mmol l(-1); 21% reduction) and the vacuolar H(+)-ATPase inhibitor bafilomycin (5 μmol l(-1); 21% reduction), and by the organic anions PAH (10 mmol l(-1); 44% reduction) and probenecid (10 mmol l(-1); 61% reduction, concentration-dependent inhibition). Transport by locust Malpighian tubules exhibits a similar pharmacological profile, albeit with markedly higher concentrations of CPR being reached in S. gregaria. Immunolocalization of the Na(+)/K(+)-ATPase α-subunit in S. gregaria revealed that this transporter is abundantly expressed and localized to the basal cell membranes. Immunolocalization data could not be obtained from H. crispae. Our results indicate that organic anion secretion by the tardigrade midgut is transporter mediated with likely candidates for the basolateral entry step being members of the Oat and/or Oatp transporter families. From our results, we cautiously suggest that apical H(+) and possibly basal Na(+)/K(+) pumps provide the driving force for the transport; the exact coupling between electrochemical gradients generated by the pumps and transport of ions, as well as the nature of the apical exit step, are unknown. This study is, to our knowledge, the first to show active epithelial transport in tardigrades.

Intracellular Requirements for Passive Proton Transport through the Na+,K+-ATPase.

PubMed

Stanley, Kevin S; Meyer, Dylan J; Gatto, Craig; Artigas, Pablo

2016-12-06

The Na + ,K + -ATPase (NKA or Na/K pump) hydrolyzes one ATP to exchange three intracellular Na+ (Na + i ) for two extracellular K+ (K + o ) across the plasma membrane by cycling through a set of reversible transitions between phosphorylated and dephosphorylated conformations, alternately opening ion-binding sites externally (E2) or internally (E1). With subsaturating [Na + ] o and [K + ] o , the phosphorylated E2P conformation passively imports protons generating an inward current (I H ), which may be exacerbated in NKA-subunit mutations associated with human disease. To elucidate the mechanisms of I H , we studied the effects of intracellular ligands (transported ions, nucleotides, and beryllium fluoride) on I H and, for comparison, on transient currents measured at normal Na + o (Q Na ). Utilizing inside-out patches from Xenopus oocytes heterologously expressing NKA, we observed that 1) in the presence of Na + i , I H and Q Na were both activated by ATP, but not ADP; 2) the [Na + ] i dependence of I H in saturating ATP showed K 0.5,Na  = 1.8 ± 0.2 mM and the [ATP] dependence at saturating [Na + ] i yielded K 0.5,ATP  = 48 ± 11 μM (in comparison, Na + i -dependent Q Na yields K 0.5,Na  = 0.8 ± 0.2 mM and K 0.5,ATP  = 0.43 ± 0.03 μM; 3) ATP activated I H in the presence of K + i (∼15% of the I H observed in Na + i ) only when Mg 2+ i was also present; and 4) beryllium fluoride induced maximal I H  even in the absence of nucleotide. These data indicate that I H occurs when NKA is in an externally open E2P state with nucleotide bound, a conformation that can be reached through forward Na/K pump phosphorylation of E1, with Na + i and ATP, or by backward binding of K + i to E1, which drives the pump to the occluded E2(2K), where free P i (at the micromolar levels found in millimolar ATP solutions) promotes external release of occluded K + by backdoor NKA phosphorylation. Maximal I H through beryllium-fluorinated NKA indicates that this complex mimics ATP

Overexpression of a novel soybean gene modulating Na+ and K+ transport enhances salt tolerance in transgenic tobacco plants.

PubMed

Chen, Huatao; He, Hui; Yu, Deyue

2011-01-01

Salt is an important factor affecting the growth and development of soybean in saline soil. In this study, a novel soybean gene encoding a transporter (GmHKT1) was identified and its function analyzed using transgenic plants. GmHKT1 encoded a protein of 419 amino acids, with a potential molecular mass of 47.06 kDa and a predicted pI value of 8.59. Comparison of the genomic and cDNA sequences of GmHKT1 identified no intron. The deduced amino acid sequence of GmHKT1 showed 38-49% identity with other plant HKT-like sequences. RT-PCR analysis showed that the expression of GmHKT1 was upregulated by salt stress (150 mM NaCl) in roots and leaves but not in stems. Overexpression of GmHKT1 significantly enhanced the tolerance of transgenic tobacco plants to salt stress, compared with non-transgenic plants. To investigate the role of GmHKT1 in K(+) and Na(+) transport, we compared K(+) and Na(+) accumulation in roots and shoots of wild-type and transgenic tobacco plants. The results suggested that GmHKT1 is a transporter that affected K(+) and Na(+) transport in roots and shoots, and regulated Na(+) /K(+) homeostasis in these organs. Our findings suggest that GmHKT1 plays an important role in response to salt stress and would be useful in engineering crop plants for enhanced tolerance to salt stress. Copyright © Physiologia Plantarum 2010.

Transport of bile acids in multidrug-resistance-protein 3-overexpressing cells co-transfected with the ileal Na+-dependent bile-acid transporter.

PubMed Central

Zelcer, Noam; Saeki, Tohru; Bot, Ilse; Kuil, Annemieke; Borst, Piet

2003-01-01

Many of the transporters involved in the transport of bile acids in the enterohepatic circulation have been characterized. The basolateral bile-acid transporter of ileocytes and cholangiocytes remains an exception. It has been suggested that rat multidrug resistance protein 3 (Mrp3) fulfills this function. Here we analyse bile-salt transport by human MRP3. Membrane vesicles from insect ( Spodoptera frugiperda ) cells expressing MRP3 show time-dependent uptake of glycocholate and taurocholate. Furthermore, sulphated bile salts were high-affinity competitive inhibitors of etoposide glucuronide transport by MRP3 (IC50 approximately 10 microM). Taurochenodeoxycholate, taurocholate and glycocholate inhibited transport at higher concentrations (IC50 approximately 100, 250 and 500 microM respectively). We used mouse fibroblast-like cell lines derived from mice with disrupted Mdr1a, Mdr1b and Mrp1 genes to generate transfectants that express the murine apical Na+-dependent bile-salt transporter (Asbt) and MRP3. Uptake of glycocholate by these cells is Na+-dependent, with a K(m) and V(max) of 29+/-7 microM and 660 +/- 63 pmol/min per mg of protein respectively and is inhibited by several organic-aniontransport inhibitors. Expression of MRP3 in these cells limits the accumulation of glycocholate and increases the efflux from cells preloaded with taurocholate or glycocholate. In conclusion, we find that MRP3 transports both taurocholate and glycocholate, albeit with low affinity, in contrast with the high-affinity transport by rat Mrp3. Our results suggest that MRP3 is unlikely to be the principal basolateral bile-acid transporter of ileocytes and cholangiocytes, but that it may have a role in the removal of bile acids from the liver in cholestasis. PMID:12220224

Transport regimes spanning magnetization-coupling phase space

DOE PAGES

Baalrud, Scott D.; Daligault, Jérôme

2017-10-06

The manner in which transport properties vary over the entire parameter-space of coupling and magnetization strength is explored in this paper. Four regimes are identified based on the relative size of the gyroradius compared to other fundamental length scales: the collision mean free path, Debye length, distance of closest approach, and interparticle spacing. Molecular dynamics simulations of self-diffusion and temperature anisotropy relaxation spanning the parameter space are found to agree well with the predicted boundaries. Finally, comparison with existing theories reveals regimes where they succeed, where they fail, and where no theory has yet been developed.

Transport regimes spanning magnetization-coupling phase space

DOE Office of Scientific and Technical Information (OSTI.GOV)

Baalrud, Scott D.; Daligault, Jérôme

The manner in which transport properties vary over the entire parameter-space of coupling and magnetization strength is explored in this paper. Four regimes are identified based on the relative size of the gyroradius compared to other fundamental length scales: the collision mean free path, Debye length, distance of closest approach, and interparticle spacing. Molecular dynamics simulations of self-diffusion and temperature anisotropy relaxation spanning the parameter space are found to agree well with the predicted boundaries. Finally, comparison with existing theories reveals regimes where they succeed, where they fail, and where no theory has yet been developed.

Single-photon transport through a waveguide coupling to a quadratic optomechanical system

NASA Astrophysics Data System (ADS)

Qiao, Lei

2017-07-01

We study the coherent transport of a single photon, which propagates in a one-dimensional waveguide and is scattered by a quadratic optomechanical system. Our approach, which is based on the Lippmann-Schwinger equation, gives an analytical solution to describe the single-photon transmission and reflection properties. We analyze the transport spectra and find they are not only related to the optomechanical system's energy-level structure, but also dependent on the optomechanical system's inherent parameters. For the existence of atomic degrees of freedom, we get a Rabi-splitting-like or an electromagnetically induced transparency (EIT)-like spectrum, depending on the atom-cavity coupling strength. Here, we focus on the single-photon strong-coupling regime so that single-quantum effects could be seen.

Coupled porohyperelastic mass transport (PHEXPT) finite element models for soft tissues using ABAQUS.

PubMed

Vande Geest, Jonathan P; Simon, B R; Rigby, Paul H; Newberg, Tyler P

2011-04-01

Finite element models (FEMs) including characteristic large deformations in highly nonlinear materials (hyperelasticity and coupled diffusive/convective transport of neutral mobile species) will allow quantitative study of in vivo tissues. Such FEMs will provide basic understanding of normal and pathological tissue responses and lead to optimization of local drug delivery strategies. We present a coupled porohyperelastic mass transport (PHEXPT) finite element approach developed using a commercially available ABAQUS finite element software. The PHEXPT transient simulations are based on sequential solution of the porohyperelastic (PHE) and mass transport (XPT) problems where an Eulerian PHE FEM is coupled to a Lagrangian XPT FEM using a custom-written FORTRAN program. The PHEXPT theoretical background is derived in the context of porous media transport theory and extended to ABAQUS finite element formulations. The essential assumptions needed in order to use ABAQUS are clearly identified in the derivation. Representative benchmark finite element simulations are provided along with analytical solutions (when appropriate). These simulations demonstrate the differences in transient and steady state responses including finite deformations, total stress, fluid pressure, relative fluid, and mobile species flux. A detailed description of important model considerations (e.g., material property functions and jump discontinuities at material interfaces) is also presented in the context of finite deformations. The ABAQUS-based PHEXPT approach enables the use of the available ABAQUS capabilities (interactive FEM mesh generation, finite element libraries, nonlinear material laws, pre- and postprocessing, etc.). PHEXPT FEMs can be used to simulate the transport of a relatively large neutral species (negligible osmotic fluid flux) in highly deformable hydrated soft tissues and tissue-engineered materials.

Vigabatrin transport across the human intestinal epithelial (Caco-2) brush-border membrane is via the H+-coupled amino-acid transporter hPAT1

PubMed Central

Abbot, Emily L; Grenade, Danielle S; Kennedy, David J; Gatfield, Kelly M; Thwaites, David T

2005-01-01

The aim of this investigation was to determine if the human proton-coupled amino-acid transporter 1 (hPAT1 or SLC36A1) is responsible for the intestinal uptake of the orally-administered antiepileptic agent 4-amino-5-hexanoic acid (vigabatrin). The Caco-2 cell line was used as a model of the human small intestinal epithelium. Competition experiments demonstrate that [3H]GABA uptake across the apical membrane was inhibited by vigabatrin and the GABA analogues trans-4-aminocrotonic acid (TACA) and guvacine, whereas 1-(aminomethyl)cyclohexaneacetic acid (gabapentin) had no affect. Experiments with 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF)-loaded Caco-2 cells demonstrate that apical exposure to vigabatrin and TACA induce comparable levels of intracellular acidification (due to H+/amino-acid symport) to that generated by GABA, suggesting that they are substrates for a H+-coupled absorptive transporter such as hPAT1. In hPAT1 and mPAT1-expressing Xenopus laevis oocytes [3H]GABA uptake was inhibited by vigabatrin, TACA and guvacine, whereas gabapentin failed to inhibit [3H]GABA uptake. In Na+-free conditions, vigabatrin and TACA evoked similar current responses (due to H+/amino-acid symport) in hPAT1-expressing oocytes under voltage-clamp conditions to that induced by GABA (whereas no current was observed in water-injected oocytes) consistent with the ability of these GABA analogues to inhibit [3H]GABA uptake. This study demonstrates that hPAT1 is the carrier responsible for the uptake of vigabatrin across the brush-border membrane of the small intestine and emphasises the therapeutic potential of hPAT1 as a delivery route for orally administered, clinically significant GABA-related compounds. PMID:16331283

L-leucine, L-methionine, and L-phenylalanine share a Na(+)/K (+)-dependent amino acid transporter in shrimp hepatopancreas.

PubMed

Duka, Ada; Ahearn, Gregory A

2013-08-01

Hepatopancreatic brush border membrane vesicles (BBMV), made from Atlantic White shrimp (Litopenaeus setiferus), were used to characterize the transport properties of (3)H-L-leucine influx by these membrane systems and how other essential amino acids and the cations, sodium and potassium, interact with this transport system. (3)H-L-leucine uptake by BBMV was pH-sensitive and occurred against transient transmembrane concentration gradients in both Na(+)- and K(+)-containing incubation media, suggesting that either cation was capable of providing a driving force for amino acid accumulation. (3)H-L-leucine uptake in NaCl or KCl media were each three times greater in acidic pH (pH 5.5) than in alkaline pH (pH 8.5). The essential amino acid, L-methionine, at 20 mM significantly (p < 0.0001) inhibited the 2-min uptakes of 1 mM (3)H-L-leucine in both Na(+)- and K(+)-containing incubation media. The residual (3)H-L-leucine uptake in the two media were significantly greater than zero (p < 0.001), but not significantly different from each other (p > 0.05) and may represent an L-methionine- and cation-independent transport system. (3)H-L-leucine influxes in both NaCl and KCl incubation media were hyperbolic functions of [L-leucine], following the carrier-mediated Michaelis-Menten equation. In NaCl, (3)H-L-leucine influx displayed a low apparent K M (high affinity) and low apparent J max, while in KCl the transport exhibited a high apparent K M (low affinity) and high apparent J max. L-methionine or L-phenylalanine (7 and 20 mM) were competitive inhibitors of (3)H-L-leucine influxes in both NaCl and KCl media, producing a significant (p < 0.01) increase in (3)H-L-leucine influx K M, but no significant response in (3)H-L-leucine influx J max. Potassium was a competitive inhibitor of sodium co-transport with (3)H-L-leucine, significantly (p < 0.01) increasing (3)H-L-leucine influx K M in the presence of sodium, but having negligible effect on (3)H-L-leucine influx J

Application of Mortar Coupling in Multiscale Modelling of Coupled Flow, Transport, and Biofilm Growth in Porous Media

NASA Astrophysics Data System (ADS)

Laleian, A.; Valocchi, A. J.; Werth, C. J.

2017-12-01

Multiscale models of reactive transport in porous media are capable of capturing complex pore-scale processes while leveraging the efficiency of continuum-scale models. In particular, porosity changes caused by biofilm development yield complex feedbacks between transport and reaction that are difficult to quantify at the continuum scale. Pore-scale models, needed to accurately resolve these dynamics, are often impractical for applications due to their computational cost. To address this challenge, we are developing a multiscale model of biofilm growth in which non-overlapping regions at pore and continuum spatial scales are coupled with a mortar method providing continuity at interfaces. We explore two decompositions of coupled pore-scale and continuum-scale regions to study biofilm growth in a transverse mixing zone. In the first decomposition, all reaction is confined to a pore-scale region extending the transverse mixing zone length. Only solute transport occurs in the surrounding continuum-scale regions. Relative to a fully pore-scale result, we find the multiscale model with this decomposition has a reduced run time and consistent result in terms of biofilm growth and solute utilization. In the second decomposition, reaction occurs in both an up-gradient pore-scale region and a down-gradient continuum-scale region. To quantify clogging, the continuum-scale model implements empirical relations between porosity and continuum-scale parameters, such as permeability and the transverse dispersion coefficient. Solutes are sufficiently mixed at the end of the pore-scale region, such that the initial reaction rate is accurately computed using averaged concentrations in the continuum-scale region. Relative to a fully pore-scale result, we find accuracy of biomass growth in the multiscale model with this decomposition improves as the interface between pore-scale and continuum-scale regions moves downgradient where transverse mixing is more fully developed. Also, this

Structural and mechanistic basis of proton-coupled metal ion transport in the SLC11/NRAMP family

PubMed Central

Ehrnstorfer, Ines A.; Manatschal, Cristina; Arnold, Fabian M.; Laederach, Juerg; Dutzler, Raimund

2017-01-01

Secondary active transporters of the SLC11/NRAMP family catalyse the uptake of iron and manganese into cells. These proteins are highly conserved across all kingdoms of life and thus likely share a common transport mechanism. Here we describe the structural and functional properties of the prokaryotic SLC11 transporter EcoDMT. Its crystal structure reveals a previously unknown outward-facing state of the protein family. In proteoliposomes EcoDMT mediates proton-coupled uptake of manganese at low micromolar concentrations. Mutants of residues in the transition-metal ion-binding site severely affect transport, whereas a mutation of a conserved histidine located near this site results in metal ion transport that appears uncoupled to proton transport. Combined with previous results, our study defines the conformational changes underlying transition-metal ion transport in the SLC11 family and it provides molecular insight to its coupling to protons. PMID:28059071

Cloning, localization and expression analysis of vacuolar sugar transporters in the CAM plant Ananas comosus (pineapple).

PubMed

Antony, Edna; Taybi, Tahar; Courbot, Mikaël; Mugford, Sam T; Smith, J Andrew C; Borland, Anne M

2008-01-01

In photosynthetic tissues of the CAM plant pineapple (Ananas comosus), storage of soluble sugars in the central vacuole during the daytime and their remobilization at night is required to provide carbon skeletons for nocturnal CO(2) fixation. However, soluble sugars produced photosynthetically must also be exported to support growth processes in heterotrophic tissues. To begin to address how vacuolar sugar storage and assimilate partitioning are regulated in A. comosus, degenerate PCR and cDNA library screening were used to clone three candidate sugar transporters from the leaves of this species. Subcellular localization of the three transporters was investigated via expression of YFP-fusion proteins in tobacco epidermal cells and their co-localization with subcellular markers by confocal microscopy. Using this strategy, a putative hexose transporter (AcMST1) and a putative inositol transporter (AcINT1) were identified that both localized to the tonoplast, whereas a putative sucrose transporter (AcSUT1) was found to localize to prevacuolar compartments. A cDNA (AcMST2) with high similarity to a recently characterized tonoplast hexose transporter in Arabidopsis was also identified from an A. comosus fruit EST database. Analyses of transcript abundance indicated that AcMST1 was more highly expressed in fruits compared to leaves of A. comosus, whilst transcripts of AcINT1, AcSUT1, and AcMST2 were more abundant in leaves. Transcript abundance of AcINT1, the putative inositol transporter, showed day-night changes comparable to those of other CAM-related transcripts described in Mesembryanthemum crystallinum. The results are discussed in terms of the role of vacuolar sugar transporters in regulating carbon flow during the diel cycle in CAM plants.

Na/K pump inactivation, subsarcolemmal Na measurements, and cytoplasmic ion turnover kinetics contradict restricted Na spaces in murine cardiac myocytes

PubMed Central

Lu, Fang-Min

2017-01-01

Decades ago, it was proposed that Na transport in cardiac myocytes is modulated by large changes in cytoplasmic Na concentration within restricted subsarcolemmal spaces. Here, we probe this hypothesis for Na/K pumps by generating constitutive transsarcolemmal Na flux with the Na channel opener veratridine in whole-cell patch-clamp recordings. Using 25 mM Na in the patch pipette, pump currents decay strongly during continuous activation by extracellular K (τ, ∼2 s). In contradiction to depletion hypotheses, the decay becomes stronger when pump currents are decreased by hyperpolarization. Na channel currents are nearly unchanged by pump activity in these conditions, and conversely, continuous Na currents up to 0.5 nA in magnitude have negligible effects on pump currents. These outcomes are even more pronounced using 50 mM Li as a cytoplasmic Na congener. Thus, the Na/K pump current decay reflects mostly an inactivation mechanism that immobilizes Na/K pump charge movements, not cytoplasmic Na depletion. When channel currents are increased beyond 1 nA, models with unrestricted subsarcolemmal diffusion accurately predict current decay (τ ∼15 s) and reversal potential shifts observed for Na, Li, and K currents through Na channels opened by veratridine, as well as for Na, K, Cs, Li, and Cl currents recorded in nystatin-permeabilized myocytes. Ion concentrations in the pipette tip (i.e., access conductance) track without appreciable delay the current changes caused by sarcolemmal ion flux. Importantly, cytoplasmic mixing volumes, calculated from current decay kinetics, increase and decrease as expected with osmolarity changes (τ >30 s). Na/K pump current run-down over 20 min reflects a failure of pumps to recover from inactivation. Simulations reveal that pump inactivation coupled with Na-activated recovery enhances the rapidity and effectivity of Na homeostasis in cardiac myocytes. In conclusion, an autoregulatory mechanism enhances cardiac Na/K pump activity when

Coupled Particle Transport and Pattern Formation in a Nonlinear Leaky-Box Model

NASA Technical Reports Server (NTRS)

Barghouty, A. F.; El-Nemr, K. W.; Baird, J. K.

2009-01-01

Effects of particle-particle coupling on particle characteristics in nonlinear leaky-box type descriptions of the acceleration and transport of energetic particles in space plasmas are examined in the framework of a simple two-particle model based on the Fokker-Planck equation in momentum space. In this model, the two particles are assumed coupled via a common nonlinear source term. In analogy with a prototypical mathematical system of diffusion-driven instability, this work demonstrates that steady-state patterns with strong dependence on the magnetic turbulence but a rather weak one on the coupled particles attributes can emerge in solutions of a nonlinearly coupled leaky-box model. The insight gained from this simple model may be of wider use and significance to nonlinearly coupled leaky-box type descriptions in general.

Inverse engineering for fast transport and spin control of spin-orbit-coupled Bose-Einstein condensates in moving harmonic traps

NASA Astrophysics Data System (ADS)

Chen, Xi; Jiang, Ruan-Lei; Li, Jing; Ban, Yue; Sherman, E. Ya.

2018-01-01

We investigate fast transport and spin manipulation of tunable spin-orbit-coupled Bose-Einstein condensates in a moving harmonic trap. Motivated by the concept of shortcuts to adiabaticity, we design inversely the time-dependent trap position and spin-orbit-coupling strength. By choosing appropriate boundary conditions we obtain fast transport and spin flip simultaneously. The nonadiabatic transport and relevant spin dynamics are illustrated with numerical examples and compared with the adiabatic transport with constant spin-orbit-coupling strength and velocity. Moreover, the influence of nonlinearity induced by interatomic interaction is discussed in terms of the Gross-Pitaevskii approach, showing the robustness of the proposed protocols. With the state-of-the-art experiments, such an inverse engineering technique paves the way for coherent control of spin-orbit-coupled Bose-Einstein condensates in harmonic traps.

Acidic pH and short-chain fatty acids activate Na+ transport but differentially modulate expression of Na+/H+ exchanger isoforms 1, 2, and 3 in omasal epithelium.

PubMed

Lu, Zhongyan; Yao, Lei; Jiang, Zhengqian; Aschenbach, Jörg R; Martens, Holger; Shen, Zanming

2016-01-01

Low sodium content in feed and large amounts of salivary sodium secretion are essential requirements to efficient sodium reabsorption in the dairy cow. It is already known that Na(+)/H(+) exchange (NHE) of the ruminal epithelium plays a key role in Na(+) absorption, and its function is influenced by the presence of short-chain fatty acids (SCFA) and mucosal pH. By contrast, the functional role and regulation of NHE in omasal epithelium have not been completely understood. In the present study, we used model studies in small ruminants (sheep and goats) to investigate NHE-mediated Na(+) transport and the effects of pH and SCFA on NHE activity in omasal epithelium and on the expression of NHE isoform in omasal epithelial cells. Conventional Ussing chamber technique, primary cell culture, quantitative PCR, and Western blot were used. In native omasal epithelium of sheep, the Na(+) transport was electroneutral, and it was inhibited by the specific NHE3 inhibitor 3-[2-(3-guanidino-2-methyl-3-oxo-propenyl)-5-methyl-phenyl]-N-isopropylidene-2-methyl-acrylamide dihydrochloride, which decreased mucosal-to-serosal, serosal-to-mucosal, and net flux rates of Na(+) by 80% each. The application of low mucosal pH (6.4 or 5.8) in the presence of SCFA activated the Na(+) transport across omasal epithelium of sheep compared with that at pH 7.4. In cultured omasal epithelial cells of goats, mRNA and protein of NHE1, NHE2, and NHE3 were detected. The application of SCFA increased NHE1 mRNA and protein expression, which was most prominent when the culture medium pH decreased from 7.4 to 6.8. At variance, the mRNA and protein expression of NHE2 and NHE3 were decreased with low pH and SCFA, which was contrary to the published data from ruminal epithelial studies. In conclusion, this paper shows that (1) NHE1, NHE2, and NHE3 are expressed in omasal epithelium; (2) NHE3 mediates the major portion of transepithelial Na(+) transport in omasal epithelium; and (3) SCFA and acidic pH acutely

The discovery of slowness: low-capacity transport and slow anion channel gating by the glutamate transporter EAAT5.

PubMed

Gameiro, Armanda; Braams, Simona; Rauen, Thomas; Grewer, Christof

2011-06-08

Excitatory amino acid transporters (EAATs) control the glutamate concentration in the synaptic cleft by glial and neuronal glutamate uptake. Uphill glutamate transport is achieved by the co-/countertransport of Na(+) and other ions down their concentration gradients. Glutamate transporters also display an anion conductance that is activated by the binding of Na(+) and glutamate but is not thermodynamically coupled to the transport process. Of the five known glutamate transporter subtypes, the retina-specific subtype EAAT5 has the largest conductance relative to glutamate uptake activity. Our results suggest that EAAT5 behaves as a slow-gated anion channel with little glutamate transport activity. At steady state, EAAT5 was activated by glutamate, with a K(m)= 61 ± 11 μM. Binding of Na(+) to the empty transporter is associated with a K(m) = 229 ± 37 mM, and binding to the glutamate-bound form is associated with a K(m) = 76 ± 40 mM. Using laser-pulse photolysis of caged glutamate, we determined the pre-steady-state kinetics of the glutamate-induced anion current of EAAT5. This was characterized by two exponential components with time constants of 30 ± 1 ms and 200 ± 15 ms, which is an order of magnitude slower than those observed in other glutamate transporters. A voltage-jump analysis of the anion currents indicates that the slow activation behavior is caused by two slow, rate-limiting steps in the transport cycle, Na(+) binding to the empty transporter, and translocation of the fully loaded transporter. We propose a kinetic transport scheme that includes these two slow steps and can account for the experimentally observed data. Overall, our results suggest that EAAT5 may not act as a classical high-capacity glutamate transporter in the retina; rather, it may function as a slow-gated glutamate receptor and/or glutamate buffering system. Copyright © 2011 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Structure/function relationships in serotonin transporter: new insights from the structure of a bacterial transporter.

PubMed

Rudnick, G

2006-01-01

Serotonin transporter (SERT) serves the important function of taking up serotonin (5-HT) released during serotonergic neurotransmission. It is the target for important therapeutic drugs and psychostimulants. SERT catalyzes the influx of 5-HT together with Na+ and Cl- in a 1:1:1 stoichiometry. In the same catalytic cycle, there is coupled efflux of one K+ ion. SERT is one member of a large family of amino acid and amine transporters that is believed to utilize similar mechanisms of transport. A bacterial member of this family was recently crystallized, revealing the structural basis of these transporters. In light of the new structure, previous results with SERT have been re-interpreted, providing new insight into the substrate binding site, the permeation pathway, and the conformational changes that occur during the transport cycle.

Structure of a pantothenate transporter and implications for ECF module sharing and energy coupling of group II ECF transporters

PubMed Central

Zhang, Minhua; Bao, Zhihao; Zhao, Qin; Guo, Hui; Xu, Ke; Wang, Chengcheng

2014-01-01

Energy-coupling factor (ECF) transporters are a unique group of ATP-binding cassette (ABC) transporters responsible for micronutrient uptake from the environment. Each ECF transporter is composed of an S component (or EcfS protein) and T/A/A′ components (or EcfT/A/A′ proteins; ECF module). Among the group II ECF transporters, several EcfS proteins share one ECF module; however, the underlying mechanism remains unknown. Here we report the structure of a group II ECF transporter–pantothenate transporter from Lactobacillus brevis (LbECF-PanT), which shares the ECF module with the folate and hydroxymethylpyrimidine transporters (LbECF-FolT and LbECF-HmpT). Structural and mutational analyses revealed the residues constituting the pantothenate-binding pocket. We found that although the three EcfS proteins PanT, FolT, and HmpT are dissimilar in sequence, they share a common surface area composed of the transmembrane helices 1/2/6 (SM1/2/6) to interact with the coupling helices 2/3 (CH2/3) of the same EcfT. CH2 interacts mainly with SM1 via hydrophobic interactions, which may modulate the sliding movement of EcfS. CH3 binds to a hydrophobic surface groove formed by SM1, SM2, and SM6, which may transmit the conformational changes from EcfA/A′ to EcfS. We also found that the residues at the intermolecular surfaces in LbECF-PanT are essential for transporter activity, and that these residues may mediate intermolecular conformational transmission and/or affect transporter complex stability. In addition, we found that the structure of EcfT is conformationally dynamic, which supports its function as a scaffold to mediate the interaction of the ECF module with various EcfS proteins to form different transporter complexes. PMID:25512487

Enhanced hexose fermentation by Saccharomyces cerevisiae through integration of stoichiometric modeling and genetic screening.

PubMed

Quarterman, Josh; Kim, Soo Rin; Kim, Pan-Jun; Jin, Yong-Su

2015-01-20

In order to determine beneficial gene deletions for ethanol production by the yeast Saccharomyces cerevisiae, we performed an in silico gene deletion experiment based on a genome-scale metabolic model. Genes coding for two oxidative phosphorylation reactions (cytochrome c oxidase and ubiquinol cytochrome c reductase) were identified by the model-based simulation as potential deletion targets for enhancing ethanol production and maintaining acceptable overall growth rate in oxygen-limited conditions. Since the two target enzymes are composed of multiple subunits, we conducted a genetic screening study to evaluate the in silico results and compare the effect of deleting various portions of the respiratory enzyme complexes. Over two-thirds of the knockout mutants identified by the in silico study did exhibit experimental behavior in qualitative agreement with model predictions, but the exceptions illustrate the limitation of using a purely stoichiometric model-based approach. Furthermore, there was a substantial quantitative variation in phenotype among the various respiration-deficient mutants that were screened in this study, and three genes encoding respiratory enzyme subunits were identified as the best knockout targets for improving hexose fermentation in microaerobic conditions. Specifically, deletion of either COX9 or QCR9 resulted in higher ethanol production rates than the parental strain by 37% and 27%, respectively, with slight growth disadvantages. Also, deletion of QCR6 led to improved ethanol production rate by 24% with no growth disadvantage. The beneficial effects of these gene deletions were consistently demonstrated in different strain backgrounds and with four common hexoses. The combination of stoichiometric modeling and genetic screening using a systematic knockout collection was useful for narrowing a large set of gene targets and identifying targets of interest. Copyright © 2014 Elsevier B.V. All rights reserved.

On degenerate coupled transport processes in porous media with memory phenomena

NASA Astrophysics Data System (ADS)

Beneš, Michal; Pažanin, Igor

2018-06-01

In this paper we prove the existence of weak solutions to degenerate parabolic systems arising from the fully coupled moisture movement, solute transport of dissolved species and heat transfer through porous materials. Physically relevant mixed Dirichlet-Neumann boundary conditions and initial conditions are considered. Existence of a global weak solution of the problem is proved by means of semidiscretization in time, proving necessary uniform estimates and by passing to the limit from discrete approximations. Degeneration occurs in the nonlinear transport coefficients which are not assumed to be bounded below and above by positive constants. Degeneracies in transport coefficients are overcome by proving suitable a-priori $L^{\\infty}$-estimates based on De Giorgi and Moser iteration technique.

Neutron Transport Models and Methods for HZETRN and Coupling to Low Energy Light Ion Transport

NASA Technical Reports Server (NTRS)

Blattnig, S.R.; Slaba, T.C.; Heinbockel, J.H.

2008-01-01

Exposure estimates inside space vehicles, surface habitats, and high altitude aircraft exposed to space radiation are highly influenced by secondary neutron production. The deterministic transport code HZETRN has been identified as a reliable and efficient tool for such studies, but improvements to the underlying transport models and numerical methods are still necessary. In this paper, the forward-backward (FB) and directionally coupled forward-backward (DC) neutron transport models are derived, numerical methods for the FB model are reviewed, and a computationally efficient numerical solution is presented for the DC model. Both models are compared to the Monte Carlo codes HETCHEDS and FLUKA, and the DC model is shown to agree closely with the Monte Carlo results. Finally, it is found in the development of either model that the decoupling of low energy neutrons from the light ion (A<4) transport procedure adversely affects low energy light ion fluence spectra and exposure quantities. A first order correction is presented to resolve the problem, and it is shown to be both accurate and efficient.

Fragmentation Pathways of Lithiated Hexose Monosaccharides

NASA Astrophysics Data System (ADS)

Abutokaikah, Maha T.; Frye, Joseph W.; Tschampel, John; Rabus, Jordan M.; Bythell, Benjamin J.

2018-05-01

We characterize the primary fragmentation reactions of three isomeric lithiated D-hexose sugars (glucose, galactose, and mannose) utilizing tandem mass spectrometry, regiospecific labeling, and theory. We provide evidence that these three isomers populate similar fragmentation pathways to produce the abundant cross-ring cleavage peaks (0,2A1 and 0,3A1). These pathways are highly consistent with the prior literature (Hofmeister et al. J. Am. Chem. Soc. 113, 5964-5970, 1991, Bythell et al. J. Am. Soc. Mass Spectrom. 28, 688-703, 2017, Rabus et al. Phys. Chem. Chem. Phys. 19, 25643-25652, 2017) and the present labeling data. However, the structure-specific energetics and rate-determining steps of these reactions differ as a function of precursor sugar and anomeric configuration. The lowest energy water loss pathways involve loss of the anomeric oxygen to furnish B1 ions. For glucose and galactose, the lithiated α-anomers generate ketone structures at C2 in a concerted reaction involving a 1,2-migration of the C2-H to the anomeric carbon (C1). In contrast, the β-anomers are predicted to form 1,3-anhydroglucose/galactose B1 ion structures. Initiation of the water loss reactions from each anomeric configuration requires distinct reactive conformers, resulting in different product ion structures. Inversion of the stereochemistry at C2 has marked consequences. Both lithiated mannose forms expel water to form 1,2-anhydromannose B1 ions with the newly formed epoxide group above the ring. Additionally, provided water loss is not instantaneous, the α-anomer can also isomerize to generate a ketone structure at C2 in a concerted reaction involving a 1,2-migration of the C2-H to C1. This product is indistinguishable to that from α-glucose. The energetics and interplay of these pathways are discussed. [Figure not available: see fulltext.

Packaging of wavelength stabilized 976nm 100W 105µm 0.15 NA fiber coupled diode lasers

NASA Astrophysics Data System (ADS)

Jiang, Xiaochen; Liu, Rui; Gao, Yanyan; Zhang, Tujia; He, Xiaoguang; Zhu, Jing; Zhang, Qiang; Yang, Thomas; Zhang, Cuipeng

2016-03-01

Fiber coupled diode lasers are widely used in many fields now especially as pumps in fiber laser systems. In many fiber laser applications, high brightness pumps are essential to achieve high brightness fiber lasers. Furthermore, 976nm wavelength absorption band is narrow with Yb3+ doped fiber lasers which is more challenging for controlling wavelength stabilized in diode laser modules. This study designed and implemented commercial available high brightness and narrow wavelength width lasers to be able to use in previous mentioned applications. Base on multiple single emitters using spatial and polarization beam combining as well as fiber coupling techniques, we report a wavelength stabilized, 105μm NA 0.15 fiber coupled diode laser package with 100W of optical output power at 976 nm, which are 14 emitters inside each multiple single emitter module. The emitting aperture of the combined lasers output are designed and optimized for coupling light into a 105μm core NA 0.15 fiber. Volume Bragg grating technology has been used to improve spectral characteristics of high-power diode lasers. Mechanical modular design and thermal simulation are carried out to optimize the package. The spectral width is roughly 0.5 nm (FWHM) and the wavelength shift per °C < 0.02nm. The output spectrum is narrowed and wavelength is stabilized using Volume Bragg gratings (VBGs). The high brightness package has an electrical to optical efficiency better than 45% and power enclosure more than 90% within NA 0.12. Qualification tests have been included on this kind of package. Mechanical shock, vibration and accelerated aging tests show that the package is reliability and the MTTF is calculated to be more than 100k hours at 25°C.

Identification and functional characterization of a Na+-independent neutral amino acid transporter with broad substrate selectivity.

PubMed

Segawa, H; Fukasawa, Y; Miyamoto, K; Takeda, E; Endou, H; Kanai, Y

1999-07-09

We have isolated a cDNA from rat small intestine that encodes a novel Na+-independent neutral amino acid transporter with distinctive characteristics in substrate selectivity and transport property. The encoded protein, designated L-type amino acid transporter-2 (LAT-2), shows amino acid sequence similarity to the system L Na+-independent neutral amino acid transporter LAT-1 (Kanai, Y., Segawa, H., Miyamoto, K., Uchino, H., Takeda, E., and Endou, H. (1998) J. Biol. Chem. 273, 23629-23632) (50% identity) and the system y+L transporters y+LAT-1 (47%) and KIAA0245/y+LAT-2 (45%) (Torrents, D., Estevez, R., Pineda, M., Fernandez, E., Lloberas, J., Shi, Y.-B., Zorzano, A., and Palacin, M. (1998) J. Biol. Chem. 273, 32437-32445). LAT-2 is a nonglycosylated membrane protein. It requires 4F2 heavy chain, a type II membrane glycoprotein, for its functional expression in Xenopus oocytes. LAT-2-mediated transport is not dependent on Na+ or Cl- and is inhibited by a system L-specific inhibitor, 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH), indicating that LAT-2 is a second isoform of the system L transporter. Compared with LAT-1, which prefers large neutral amino acids with branched or aromatic side chains, LAT-2 exhibits remarkably broad substrate selectivity. It transports all of the L-isomers of neutral alpha-amino acids. LAT-2 exhibits higher affinity (Km = 30-50 microM) to Tyr, Phe, Trp, Thr, Asn, Ile, Cys, Ser, Leu, Val, and Gln and relatively lower affinity (Km = 180-300 microM) to His, Ala, Met, and Gly. In addition, LAT-2 mediates facilitated diffusion of substrate amino acids, as distinct from LAT-1, which mediates amino acid exchange. LAT-2-mediated transport is increased by lowering the pH level, with peak activity at pH 6.25, because of the decrease in the Km value without changing the Vmax value. Because of these functional properties and a high level of expression of LAT-2 in the small intestine, kidney, placenta, and brain, it is suggested that the

Protein Phosphatase 2A Interacts with the Na+,K+-ATPase and Modulates Its Trafficking by Inhibition of Its Association with Arrestin

PubMed Central

Kimura, Toru; Han, WonSun; Pagel, Philipp; Nairn, Angus C.; Caplan, Michael J.

2011-01-01

Background The P-type ATPase family constitutes a collection of ion pumps that form phosphorylated intermediates during ion transport. One of the best known members of this family is the Na+,K+-ATPase. The catalytic subunit of the Na+,K+-ATPase includes several functional domains that determine its enzymatic and trafficking properties. Methodology/Principal Findings Using the yeast two-hybrid system we found that protein phosphatase 2A (PP2A) catalytic C-subunit is a specific Na+,K+-ATPase interacting protein. PP-2A C-subunit interacted with the Na+,K+-ATPase, but not with the homologous sequences of the H+,K+-ATPase. We confirmed that the Na+,K+-ATPase interacts with a complex of A- and C-subunits in native rat kidney. Arrestins and G-protein coupled receptor kinases (GRKs) are important regulators of G-protein coupled receptor (GPCR) signaling, and they also regulate Na+,K+-ATPase trafficking through direct association. PP2A inhibits association between the Na+,K+-ATPase and arrestin, and diminishes the effect of arrestin on Na+,K+-ATPase trafficking. GRK phosphorylates the Na+,K+-ATPase and PP2A can at least partially reverse this phosphorylation. Conclusions/Significance Taken together, these data demonstrate that the sodium pump belongs to a growing list of ion transport proteins that are regulated through direct interactions with the catalytic subunit of a protein phosphatase. PMID:22242112

Parasympathetic control of Na, K transport in perfused submaxillary duct of the rat.

PubMed

Schneyer, L H

1977-07-01

The effects of stimulating the parasympathetic innervation to rat submaxillary gland on ductal transport of Na, K, water, and transepithelial PD, were tested in the main excretory duct during perfusion through its lumen. During nerve stimulation, transepithelial PD was consistently decreased, usually by about 15 mV, and this decrease could be blocked by atropine but not by adrenergic blocking agents. Net flux of Na, K, or water was not significantly changed during stimulation. One-way flux of Na from the lumen also was unaffected. The decrease in PD during stimulation was not affected by substituting isethionate for Cl or increasing [K] in the luminal perfusion medium. It is concluded that parasympathetic effects are exerted on the ductal cells, but that these probably do not involve conductance changes at the luminal membrane.

A COMSOL-GEMS interface for modeling coupled reactive-transport geochemical processes

NASA Astrophysics Data System (ADS)

Azad, Vahid Jafari; Li, Chang; Verba, Circe; Ideker, Jason H.; Isgor, O. Burkan

2016-07-01

An interface was developed between COMSOL MultiphysicsTM finite element analysis software and (geo)chemical modeling platform, GEMS, for the reactive-transport modeling of (geo)chemical processes in variably saturated porous media. The two standalone software packages are managed from the interface that uses a non-iterative operator splitting technique to couple the transport (COMSOL) and reaction (GEMS) processes. The interface allows modeling media with complex chemistry (e.g. cement) using GEMS thermodynamic database formats. Benchmark comparisons show that the developed interface can be used to predict a variety of reactive-transport processes accurately. The full functionality of the interface was demonstrated to model transport processes, governed by extended Nernst-Plank equation, in Class H Portland cement samples in high pressure and temperature autoclaves simulating systems that are used to store captured carbon dioxide (CO2) in geological reservoirs.

Structure and function of SemiSWEET and SWEET sugar transporters.

PubMed

Feng, Liang; Frommer, Wolf B

2015-08-01

SemiSWEETs and SWEETs have emerged as unique sugar transporters. First discovered in plants with the help of fluorescent biosensors, homologs exist in all kingdoms of life. Bacterial and plant homologs transport hexoses and sucrose, whereas animal SWEETs transport glucose. Prokaryotic SemiSWEETs are small and comprise a parallel homodimer of an approximately 100 amino acid-long triple helix bundle (THB). Duplicated THBs are fused to create eukaryotic SWEETs in a parallel orientation via an inversion linker helix, producing a similar configuration to that of SemiSWEET dimers. Structures of four SemiSWEETs have been resolved in three states: open outside, occluded, and open inside, indicating alternating access. As we discuss here, these atomic structures provide a basis for exploring the evolution of structure-function relations in this new class of transporters. Copyright © 2015 Elsevier Ltd. All rights reserved.

Phenformin and 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR) activation of AMP-activated protein kinase inhibits transepithelial Na+ transport across H441 lung cells.

PubMed

Woollhead, Alison M; Scott, John W; Hardie, D Grahame; Baines, Deborah L

2005-08-01

Active re-absorption of Na+ across the alveolar epithelium is essential to maintain lung fluid balance. Na+ entry at the luminal membrane is predominantly via the amiloride-sensitive Na+ channel (ENaC) down its electrochemical gradient. This gradient is generated and maintained by basolateral Na+ extrusion via Na+,K+-ATPase an energy-dependent process. Several kinases and factors that activate them are known to regulate these processes; however, the role of AMP-activated protein kinase (AMPK) in the lung is unknown. AMPK is an ultra-sensitive cellular energy sensor that monitors energy consumption and down-regulates ATP-consuming processes when activated. The biguanide phenformin has been shown to independently decrease ion transport processes, influence cellular metabolism and activate AMPK. The AMP mimetic drug 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR) also activates AMPK in intact cells. Western blotting revealed that both the alpha1 and alpha2 catalytic subunits of AMPK are present in Na+ transporting H441 human lung epithelial cells. Phenformin and AICAR increased AMPK activity in H441 cells in a dose-dependent fashion, stimulating the kinase maximally at 5-10 mm (P = 0.001, n = 3) and 2 mm (P < 0.005, n = 3), respectively. Both agents significantly decreased basal ion transport (measured as short circuit current) across H441 monolayers by approximately 50% compared with that of controls (P < 0.05, n = 4). Neither treatment altered the resistance of the monolayers. Phenformin and AICAR significantly reduced amiloride-sensitive transepithelial Na+ transport compared with controls (P < 0.05, n = 4). This was a result of both decreased Na+,K+-ATPase activity and amiloride-sensitive apical Na+ conductance. Transepithelial Na+ transport decreased with increasing concentrations of phenformin (0.1-10 mm) and showed a significant correlation with AMPK activity. Taken together, these results show that phenformin and AICAR suppress amiloride

The quick and ultrasensitive determination of K in NaI using inductively coupled plasma mass spectrometry

DOE Office of Scientific and Technical Information (OSTI.GOV)

Arnquist, Isaac J.; Hoppe, Eric W.

A highly sensitive, novel and quick assay method utilizing inductively coupled plasma mass spectrometry was developed for the determination of K in NaI powders and NaI(Tl) scintillator crystals for use in ultralow background applications. The determination of K (viz. 40K), as well as Th and U and their daughters, is important in ultralow background detector materials to ensure incorporation of materials of sufficiently high radiopurity. Through the use of improved instrumentation, cool plasma operating conditions, and meticulously clean sample preparations, detection limits of 11 fg natK∙g-1 (or 341 pBq 40K∙kg-1) was attained for K in pure water. Detection limits inmore » the sample matrix (i.e., NaI) were 0.529 ng natK∙g NaI-1 (or 16.4 Bq 40K∙kg NaI -1). A number of different precursor NaI powder samples and NaI(Tl) scintillator crystals were assayed for their K content. Determinations ranged from 0.757 – 31.4 ng natK∙g NaI-1. This method allows for the screening of materials to unprecedented levels in a fraction of the time compared to gamma counting techniques, providing a useful method for a more effective screening tool of K in ultralow background detector materials.« less

Transport Regimes Spanning Magnetization-Coupling Phase Space

NASA Astrophysics Data System (ADS)

Baalrud, Scott D.; Tiwari, Sanat; Daligault, Jerome

2017-10-01

The manner in which transport properties vary over the entire parameter-space of coupling and magnetization strength is explored. Four regimes are identified based on the relative size of the gyroradius compared to other fundamental length scales: the collision mean free path, Debye length, distance of closest approach and interparticle spacing. Molecular dynamics simulations of self-diffusion and temperature anisotropy relaxation spanning the parameter space are found to agree well with the predicted boundaries. Comparison with existing theories reveals regimes where they succeed, where they fail, and where no theory has yet been developed. The results suggest that magnetic fields may be used to assist ultracold neutral plasma experiments to reach regimes of stronger electron coupling by reducing heating of electrons in the direction perpendicular to the magnetic field.. By constraining electron motion along the direction of the magnetic field, the overall electron temperature is reduced nearly by a factor of three. A large temperature anisotropy develops as a result, which can be maintained for a long time in the regime of high electron magnetization. Work supported by LDRD project 20150520ER at LANL, AFOSR FA9550-16-1-0221 and US DOE Award DE-SC00161.

Hexose Oxidase-Mediated Hydrogen Peroxide as a Mechanism for the Antibacterial Activity in the Red Seaweed Ptilophora subcostata

PubMed Central

Ogasawara, Kimi; Yamada, Kenji; Hatsugai, Noriyuki; Imada, Chiaki; Nishimura, Mikio

2016-01-01

Marine algae have unique defense strategies against microbial infection. However, their mechanisms of immunity remain to be elucidated and little is known about the similarity of the immune systems of marine algae and terrestrial higher plants. Here, we suggest a possible mechanism underlying algal immunity, which involves hexose oxidase (HOX)-dependent production of hydrogen peroxide (H2O2). We examined crude extracts from five different red algal species for their ability to prevent bacterial growth. The extract from one of these algae, Ptilophora subcostata, was particularly active and prevented the growth of gram-positive and -negative bacteria, which was completely inhibited by treatment with catalase. The extract did not affect the growth of either a yeast or a filamentous fungus. We partially purified from P. subcostata an enzyme involved in its antibacterial activity, which shared 50% homology with the HOX of red seaweed Chondrus crispus. In-gel carbohydrate oxidase assays revealed that P. subcostata extract had the ability to produce H2O2 in a hexose-dependent manner and this activity was highest in the presence of galactose. In addition, Bacillus subtilis growth was strongly suppressed near P. subcostata algal fronds on GYP agar plates. These results suggest that HOX plays a role in P. subcostata resistance to bacterial attack by mediating H2O2 production in the marine environment. PMID:26867214

Molecular Cloning and Functional Analysis of a Na+-Insensitive K+ Transporter of Capsicum chinense Jacq

PubMed Central

Ruiz-Lau, Nancy; Bojórquez-Quintal, Emanuel; Benito, Begoña; Echevarría-Machado, Ileana; Sánchez-Cach, Lucila A.; Medina-Lara, María de Fátima; Martínez-Estévez, Manuel

2016-01-01

High-affinity K+ (HAK) transporters are encoded by a large family of genes and are ubiquitous in the plant kingdom. These HAK-type transporters participate in low- and high-affinity potassium (K+) uptake and are crucial for the maintenance of K+ homeostasis under hostile conditions. In this study, the full-length cDNA of CcHAK1 gene was isolated from roots of the habanero pepper (Capsicum chinense). CcHAK1 expression was positively regulated by K+ starvation in roots and was not inhibited in the presence of NaCl. Phylogenetic analysis placed the CcHAK1 transporter in group I of the HAK K+ transporters, showing that it is closely related to Capsicum annuum CaHAK1 and Solanum lycopersicum LeHAK5. Characterization of the protein in a yeast mutant deficient in high-affinity K+ uptake (WΔ3) suggested that CcHAK1 function is associated with high-affinity K+ uptake, with Km and Vmax for Rb of 50 μM and 0.52 nmol mg−1 min−1, respectively. K+ uptake in yeast expressing the CcHAK1 transporter was inhibited by millimolar concentrations of the cations ammonium (NH4+) and cesium (Cs+) but not by sodium (Na+). The results presented in this study suggest that the CcHAK1 transporter may contribute to the maintenance of K+ homeostasis in root cells in C. chinense plants undergoing K+-deficiency and salt stress. PMID:28083010

AMP-activated protein kinase (AMPK)-dependent and -independent pathways regulate hypoxic inhibition of transepithelial Na+ transport across human airway epithelial cells.

PubMed

Tan, C D; Smolenski, R T; Harhun, M I; Patel, H K; Ahmed, S G; Wanisch, K; Yáñez-Muñoz, R J; Baines, D L

2012-09-01

Pulmonary transepithelial Na(+) transport is reduced by hypoxia, but in the airway the regulatory mechanisms remain unclear. We investigated the role of AMPK and ROS in the hypoxic regulation of apical amiloride-sensitive Na(+) channels and basolateral Na(+) K(+) ATPase activity. H441 human airway epithelial cells were used to examine the effects of hypoxia on Na(+) transport, AMP : ATP ratio and AMPK activity. Lentiviral constructs were used to modify cellular AMPK abundance and activity; pharmacological agents were used to modify cellular ROS. AMPK was activated by exposure to 3% or 0.2% O(2) for 60 min in cells grown in submerged culture or when fluid (0.1 mL·cm(-2) ) was added to the apical surface of cells grown at the air-liquid interface. Only 0.2% O(2) activated AMPK in cells grown at the air-liquid interface. AMPK activation was associated with elevation of cellular AMP:ATP ratio and activity of the upstream kinase LKB1. Hypoxia inhibited basolateral ouabain-sensitive I(sc) (I(ouabain) ) and apical amiloride-sensitive Na(+) conductance (G(Na+) ). Modification of AMPK activity prevented the effect of hypoxia on I(ouabain) (Na(+) K(+) ATPase) but not apical G(Na+) . Scavenging of superoxide and inhibition of NADPH oxidase prevented the effect of hypoxia on apical G(Na+) (epithelial Na(+) channels). Hypoxia activates AMPK-dependent and -independent pathways in airway epithelial cells. Importantly, these pathways differentially regulate apical Na(+) channels and basolateral Na(+) K(+) ATPase activity to decrease transepithelial Na(+) transport. Luminal fluid potentiated the effect of hypoxia and activated AMPK, which could have important consequences in lung disease conditions. © 2012 The Authors. British Journal of Pharmacology © 2012 The British Pharmacological Society.

Generic Procedure for Coupling the PHREEQC Geochemical Modeling Framework with Flow and Solute Transport Simulators

NASA Astrophysics Data System (ADS)

Wissmeier, L. C.; Barry, D. A.

2009-12-01

Computer simulations of water availability and quality play an important role in state-of-the-art water resources management. However, many of the most utilized software programs focus either on physical flow and transport phenomena (e.g., MODFLOW, MT3DMS, FEFLOW, HYDRUS) or on geochemical reactions (e.g., MINTEQ, PHREEQC, CHESS, ORCHESTRA). In recent years, several couplings between both genres of programs evolved in order to consider interactions between flow and biogeochemical reactivity (e.g., HP1, PHWAT). Software coupling procedures can be categorized as ‘close couplings’, where programs pass information via the memory stack at runtime, and ‘remote couplings’, where the information is exchanged at each time step via input/output files. The former generally involves modifications of software codes and therefore expert programming skills are required. We present a generic recipe for remotely coupling the PHREEQC geochemical modeling framework and flow and solute transport (FST) simulators. The iterative scheme relies on operator splitting with continuous re-initialization of PHREEQC and the FST of choice at each time step. Since PHREEQC calculates the geochemistry of aqueous solutions in contact with soil minerals, the procedure is primarily designed for couplings to FST’s for liquid phase flow in natural environments. It requires the accessibility of initial conditions and numerical parameters such as time and space discretization in the input text file for the FST and control of the FST via commands to the operating system (batch on Windows; bash/shell on Unix/Linux). The coupling procedure is based on PHREEQC’s capability to save the state of a simulation with all solid, liquid and gaseous species as a PHREEQC input file by making use of the dump file option in the TRANSPORT keyword. The output from one reaction calculation step is therefore reused as input for the following reaction step where changes in element amounts due to advection

Measurement of mitochondrial Ca2+ transport mediated by three transport proteins: VDAC1, the Na+/Ca2+ exchanger, and the Ca2+ uniporter.

PubMed

Ben-Hail, Danya; Palty, Raz; Shoshan-Barmatz, Varda

2014-02-01

Ca(2+) is a ubiquitous cellular signal, with changes in intracellular Ca(2+) concentration not only stimulating a number of intercellular events but also triggering cell death pathways, including apoptosis. Mitochondrial Ca(2+) uptake and release play pivotal roles in cellular physiology by regulating intracellular Ca(2+) signaling, energy metabolism and cell death. Ca(2+) transport across the inner and outer mitochondrial membranes is mediated by several proteins, including channels, antiporters, and a uniporter. In this article, we present the background to several methods now established for assaying mitochondrial Ca(2+) transport activity across both mitochondrial membranes. The first of these is Ca(2+) transport mediated by the outer mitochondrial protein, the voltage-dependent anion-selective channel protein 1 (VDAC1, also known as porin 1), both as a purified protein reconstituted into a planar lipid bilayer (PLB) or into liposomes and as a mitochondrial membrane-embedded protein. The second method involves isolated mitochondria for assaying the activity of an inner mitochondrial membrane transport protein, the mitochondrial Ca(2+) uniporter (MCU) that transports Ca(2+) and is powered by the steep mitochondrial membrane potential. In the event of Ca(2+) overload, this leads to opening of the mitochondrial permeability transition pore (MPTP) and cell death. The third method describes how Na(+)-dependent mitochondrial Ca(2+) efflux mediated by mitochondrial NCLX, a member of the Na(+)/Ca(2+) exchanger superfamily, can be assayed in digitonin-permeabilized HEK-293 cells. The Ca(2+)-transport assays can be performed under various conditions and in combination with inhibitors, allowing detailed characterization of the transport activity of interest.

Biochemistry and physiology of hexose-6-phosphate knockout mice.

PubMed

Zielinska, Agnieszka E; Walker, Elizabeth A; Stewart, Paul M; Lavery, Gareth G

2011-04-10

Hexose-6-phosphate dehydrogenase (H6PDH) has emerged as an important factor in setting the redox status of the endoplasmic reticulum (ER) lumen. An important role of H6PDH is to generate a high NADPH/NADP(+) ratio which permits 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) to act as an oxo-reductase, catalyzing the activation of glucocorticoids (GCs). In H6PDH knockout mice 11β-HSD1 assumes dehydrogenase activity and inactivates GCs, rendering the target cell relatively GC insensitive. Consequently, H6PDHKO mice have a phenotype consistent with defects in the permissive and adaptive actions of GCs upon physiology. H6PDHKO mice have also offered an insight into muscle physiology as they also present with a severe vacuolating myopathy, abnormalities of glucose homeostasis and activation of the unfolded protein response due to ER stress, and a number of mechanisms driving this phenotype are thought to be involved. This article will review what we understand of the redox control of GC hormone metabolism regulated by H6PDH, and how H6PDHKO mice have allowed an in-depth understanding of its potentially novel, GC-independent roles in muscle physiology. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

Substituted cysteine accessibility method (SCAM) analysis of the transport domain of human concentrative nucleoside transporter 3 (hCNT3) and other family members reveals features of structural and functional importance

PubMed Central

Mulinta, Ras; Yao, Sylvia Y. M.; Ng, Amy M. L.; Cass, Carol E.; Young, James D.

2017-01-01

The human SLC28 family of concentrative nucleoside transporter (CNT) proteins has three members: hCNT1, hCNT2, and hCNT3. Na+-coupled hCNT1 and hCNT2 transport pyrimidine and purine nucleosides, respectively, whereas hCNT3 transports both pyrimidine and purine nucleosides utilizing Na+ and/or H+ electrochemical gradients. Escherichia coli CNT family member NupC resembles hCNT1 in permeant selectivity but is H+-coupled. Using heterologous expression in Xenopus oocytes and the engineered cysteine-less hCNT3 protein hCNT3(C−), substituted cysteine accessibility method analysis with the membrane-impermeant thiol reactive reagent p-chloromercuribenzene sulfonate was performed on the transport domain (interfacial helix 2, hairpin 1, putative transmembrane domain (TM) 7, and TM8), as well as TM9 of the scaffold domain of the protein. This systematic scan of the entire C-terminal half of hCNT3(C−) together with parallel studies of the transport domain of wild-type hCNT1 and the corresponding TMs of cysteine-less NupC(C−) yielded results that validate the newly developed structural homology model of CNT membrane architecture for human CNTs, revealed extended conformationally mobile regions within transport-domain TMs, identified pore-lining residues of functional importance, and provided evidence of an emerging novel elevator-type mechanism of transporter function. PMID:28385889

Cross-talk between ATP-regulated K+ channels and Na+ transport via cellular metabolism in frog skin principal cells.

PubMed Central

Urbach, V; Van Kerkhove, E; Maguire, D; Harvey, B J

1996-01-01

Isolated frog skin epithelium, mounted in an Ussing chamber and bathed in standard NaCl Ringer solution, recycles K+ across the basolateral membrane of principal cells through an inward-rectifier K+ channel (Kir) operating in parallel with a Na+-K+-ATPase pump. Here we report on the metabolic control of the Kir channel using patch clamping, short-circuit current measurement and enzymatic determination of cellular (ATP (ATPi). 2. The constitutively active Kir channel in the basolateral membrane has the characteristics of an ATP-regulated K+ channel and is now classed as a KATP channel. In excised inside-out patches the open probability (Po) of KATP channels was reduced by ATPi with half-maximum inhibition at an ATPi concentration of 50 microM. 3. ATPi measured (under normal Na+ transport conditions) with luciferin-luciferase was 1.50 +/- 0.23 mM (mean +/- S.E.M.; range, 0.4-3.3 mM n = 11). Thus the KATP channel would be expected to be inactive in intact cells if ATPi was the sole regulator of channel activity. KATP channels which were inactivated by 1 mM ATPi in excised patches could be reactivated by addition of 100 microM ADP on the cytosolic side. When added alone, ADP blocks this channel with half-maximal inhibition at [ADPi] > 5 mM. 4. Sulphonylureas inhibit single KATP channels in cell-attached patches as well as the total basolateral K+ current measured in frog skin epithelia perforated with nystatin on the apical side. 5. Na+-K+-ATPase activity is a major determinant of cytosolic ATP. Blocking the pump activity with ouabain produced a time-dependent increase in ATPi and reduced the open probability of KATP channels in cell-attached membranes. 6. We conclude that the ratio of ATP/ADP is an important metabolic coupling factor between the rate of Na+-K+ pumping and K+ recycling. Images Figure 9 PMID:9011625

Phenformin and 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR) activation of AMP-activated protein kinase inhibits transepithelial Na+ transport across H441 lung cells

PubMed Central

Woollhead, Alison M; Scott, John W; Hardie, D Grahame; Baines, Deborah L

2005-01-01

Active re-absorption of Na+ across the alveolar epithelium is essential to maintain lung fluid balance. Na+ entry at the luminal membrane is predominantly via the amiloride-sensitive Na+ channel (ENaC) down its electrochemical gradient. This gradient is generated and maintained by basolateral Na+ extrusion via Na+,K+-ATPase an energy-dependent process. Several kinases and factors that activate them are known to regulate these processes; however, the role of AMP-activated protein kinase (AMPK) in the lung is unknown. AMPK is an ultra-sensitive cellular energy sensor that monitors energy consumption and down-regulates ATP-consuming processes when activated. The biguanide phenformin has been shown to independently decrease ion transport processes, influence cellular metabolism and activate AMPK. The AMP mimetic drug 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR) also activates AMPK in intact cells. Western blotting revealed that both the α1 and α2 catalytic subunits of AMPK are present in Na+ transporting H441 human lung epithelial cells. Phenformin and AICAR increased AMPK activity in H441 cells in a dose-dependent fashion, stimulating the kinase maximally at 5–10 mm (P = 0.001, n = 3) and 2 mm (P < 0.005, n = 3), respectively. Both agents significantly decreased basal ion transport (measured as short circuit current) across H441 monolayers by approximately 50% compared with that of controls (P < 0.05, n = 4). Neither treatment altered the resistance of the monolayers. Phenformin and AICAR significantly reduced amiloride-sensitive transepithelial Na+ transport compared with controls (P < 0.05, n = 4). This was a result of both decreased Na+,K+-ATPase activity and amiloride-sensitive apical Na+ conductance. Transepithelial Na+ transport decreased with increasing concentrations of phenformin (0.1–10 mm) and showed a significant correlation with AMPK activity. Taken together, these results show that phenformin and AICAR suppress amiloride

ZxNHX controls Na+ and K+ homeostasis at the whole-plant level in Zygophyllum xanthoxylum through feedback regulation of the expression of genes involved in their transport

PubMed Central

Yuan, Hui-Jun; Ma, Qing; Wu, Guo-Qiang; Wang, Pei; Hu, Jing; Wang, Suo-Min

2015-01-01

Background and Aims In order to cope with arid environments, the xerohalophyte Zygophyllum xanthoxylum efficiently compartmentalizes Na+ into vacuoles, mediated by ZxNHX, and maintains stability of K+ in its leaves. However, the function of ZxNHX in controlling Na+ and K+ homeostasis at the whole-plant level remains unclear. In this study, the role of ZxNHX in regulating the expression of genes involved in Na+ and K+ transport and spatial distribution was investigated. Methods The role of ZxNHX in maintaining Na+ and K+ homeostasis in Z. xanthoxylum was studied using post-transcriptional gene silencing via  Agrobacterium-mediated transformation. Transformed plants were grown with or without 50 mm NaCl, and expression levels and physiological parameters were measured. Key Results It was found that 50 mm NaCl induced a 620 % increase in transcripts of ZxSOS1 but only an 80 % increase in transcripts of ZxHKT1;1 in roots of wild-type (WT) plants. Consequently, the ability of ZxSOS1 to transport Na+ exceeded that of ZxHKT1;1, and Na+ was loaded into the xylem by ZxSOS1 and delivered to the shoots. However, in a ZxNHX-silenced line (L7), the capacity to sequester Na+ into vacuoles of leaves was weakened, which in turn regulated long-distance Na+ transport from roots to shoots. In roots of L7, NaCl (50 mm) increased transcripts of ZxSOS1 by only 10 %, whereas transcripts of ZxHKT1;1 increased by 53 %. Thus, in L7, the transport ability of ZxHKT1;1 for Na+ outweighed that of ZxSOS1. Na+ was unloaded from the xylem stream, consequently reducing Na+ accumulation and relative distribution in leaves, but increasing the relative distribution of Na+ in roots and the net selective transport capacity for K+ over Na+ from roots to shoots compared with the WT. Silencing of ZxNHX also triggered a downregulation of  ZxAKT1 and ZxSKOR in roots, resulting in a significant decrease in K+ accumulation in all the tissues in plants grown in 50 mm NaCl. These changes led to a significant

TReacLab: An object-oriented implementation of non-intrusive splitting methods to couple independent transport and geochemical software

NASA Astrophysics Data System (ADS)

Jara, Daniel; de Dreuzy, Jean-Raynald; Cochepin, Benoit

2017-12-01

Reactive transport modeling contributes to understand geophysical and geochemical processes in subsurface environments. Operator splitting methods have been proposed as non-intrusive coupling techniques that optimize the use of existing chemistry and transport codes. In this spirit, we propose a coupler relying on external geochemical and transport codes with appropriate operator segmentation that enables possible developments of additional splitting methods. We provide an object-oriented implementation in TReacLab developed in the MATLAB environment in a free open source frame with an accessible repository. TReacLab contains classical coupling methods, template interfaces and calling functions for two classical transport and reactive software (PHREEQC and COMSOL). It is tested on four classical benchmarks with homogeneous and heterogeneous reactions at equilibrium or kinetically-controlled. We show that full decoupling to the implementation level has a cost in terms of accuracy compared to more integrated and optimized codes. Use of non-intrusive implementations like TReacLab are still justified for coupling independent transport and chemical software at a minimal development effort but should be systematically and carefully assessed.

Ifosfamide metabolites CAA, 4-OH-Ifo and Ifo-mustard reduce apical phosphate transport by changing NaPi-IIa in OK cells.

PubMed

Patzer, L; Hernando, N; Ziegler, U; Beck-Schimmer, B; Biber, J; Murer, H

2006-11-01

Renal Fanconi syndrome occurs in about 1-5% of all children treated with Ifosfamide (Ifo) and impairment of renal phosphate reabsorption in about 20-30% of them. Pathophysiological mechanisms of Ifo-induced nephropathy are ill defined. The aim has been to investigate whether Ifo metabolites affect the type IIa sodium-dependent phosphate transporter (NaPi-IIa) in viable opossum kidney cells. Ifo did not influence viability of cells or NaPi-IIa-mediated transport up to 1 mM/24 h. Incubation of confluent cells with chloroacetaldehyde (CAA) and 4-hydroperoxyIfosfamide (4-OH-Ifo) led to cell death by necrosis in a concentration-dependent manner. At low concentrations (50-100 microM/24 h), cell viability was normal but apical phosphate transport, NaPi-IIa protein, and -mRNA expression were significantly reduced. Coincubation with sodium-2-mercaptoethanesulfonate (MESNA) prevented the inhibitory action of CAA but not of 4-OH-Ifo; DiMESNA had no effect. Incubation with Ifosfamide-mustard (Ifo-mustard) did alter cell viability at concentrations above 500 microM/24 h. At lower concentrations (50-100 microM/24 h), it led to significant reduction in phosphate transport, NaPi-IIa protein, and mRNA expression. MESNA did not block these effects. The effect of Ifo-mustard was due to internalization of NaPi-IIa. Cyclophosphamide-mustard (CyP-mustard) did not have any influence on cell survival up to 1000 microM, but the inhibitory effect on phosphate transport and on NaPi-IIa protein was the same as found after Ifo-mustard. In conclusion, CAA, 4-OH-Ifo, and Ifo- and CyP-mustard are able to inhibit sodium-dependent phosphate cotransport in viable opossum kidney cells. The Ifo-mustard effect took place via internalization and reduction of de novo synthesis of NaPi-IIa. Therefore, it is possible that Ifo-mustard plays an important role in pathogenesis of Ifo-induced nephropathy.

Experimental validation of a coupled neutron-photon inverse radiation transport solver

NASA Astrophysics Data System (ADS)

Mattingly, John; Mitchell, Dean J.; Harding, Lee T.

2011-10-01

Sandia National Laboratories has developed an inverse radiation transport solver that applies nonlinear regression to coupled neutron-photon deterministic transport models. The inverse solver uses nonlinear regression to fit a radiation transport model to gamma spectrometry and neutron multiplicity counting measurements. The subject of this paper is the experimental validation of that solver. This paper describes a series of experiments conducted with a 4.5 kg sphere of α-phase, weapons-grade plutonium. The source was measured bare and reflected by high-density polyethylene (HDPE) spherical shells with total thicknesses between 1.27 and 15.24 cm. Neutron and photon emissions from the source were measured using three instruments: a gross neutron counter, a portable neutron multiplicity counter, and a high-resolution gamma spectrometer. These measurements were used as input to the inverse radiation transport solver to evaluate the solver's ability to correctly infer the configuration of the source from its measured radiation signatures.

Light-driven solute transport in Halobacterium halobium

NASA Technical Reports Server (NTRS)

Lanyi, J. K.

1979-01-01

The cell membrane of Halobacterium halobium exhibits differential regions which contain crystalline arrays of a single kind of protein, termed bacteriorhodopsin. This bacterial retinal-protein complex resembles the visual pigment and, after the absorption of protons, translocates H(+) across the cell membrane, leading to an electrochemical gradient for protons between the inside and the outside of the cell. Thus, light is an alternate source of energy in these bacteria, in addition to terminal oxidation. The paper deals with work on light-driven transport in H. halobium with cell envelope vesicles. The discussion covers light-driven movements of H(+), Na(+), and K(+); light-driven amino acid transport; and apparent allosteric control of amino acid transport. The scheme of energy coupling in H. halobium vesicles appears simple, its quantitative details are quite complex and reveal regulatory phenomena. More knowledge is required of the way the coupling components are regulated by the ion gradients present.

Coupled Eulerian-Lagrangian transport of large debris by tsunamis

NASA Astrophysics Data System (ADS)

Conde, Daniel A. S.; Ferreira, Rui M. L.; Sousa Oliveira, Carlos

2016-04-01

Tsunamis are notorious for the large disruption they can cause on coastal environments, not only due to the imparted momentum of the incoming wave but also due to its capacity to transport large quantities of solid debris, either from natural or human-made sources, over great distances. A 2DH numerical model under development at CERIS-IST (Ferreira et al., 2009; Conde, 2013) - STAV2D - capable of simulating solid transport in both Eulerian and Lagrangian paradigms will be used to assess the relevance of Lagrangian-Eulerian coupling when modelling the transport of solid debris by tsunamis. The model has been previously validated and applied to tsunami scenarios (Conde, 2013), being well-suited for overland tsunami propagation and capable of handling morphodynamic changes in estuaries and seashores. The discretization scheme is an explicit Finite Volume technique employing flux-vector splitting and a reviewed Roe-Riemann solver. Source term formulations are employed in a semi-implicit way, including the two-way coupling of the Lagrangian and Eulerian solvers by means of conservative mass and momentum transfers between fluid and solid phases. The model was applied to Sines Port, a major commercial port in Portugal, where two tsunamigenic scenarios are considered: an 8.5 Mw scenario, consistent with the Great Lisbon Earthquake and Tsunami of the 1st November 1755 (Baptista, 2009), and an hypothetical 9.5 Mw worst-case scenario based on the same historical event. Open-ocean propagation of these scenarios were simulated with GeoClaw model from ClawPack (Leveque, 2011). Following previous efforts on the modelling of debris transport by tsunamis in seaports (Conde, 2015), this work discusses the sensitivity of the obtained results with respect to the phenomenological detail of the employed Eulerian-Lagrangian formulation and the resolution of the mesh used in the Eulerian solver. The results have shown that the fluid to debris mass ratio is the key parameter regarding the

Identification of the coupling step in Na(+)-translocating NADH:quinone oxidoreductase from real-time kinetics of electron transfer.

PubMed

Belevich, Nikolai P; Bertsova, Yulia V; Verkhovskaya, Marina L; Baykov, Alexander A; Bogachev, Alexander V

2016-02-01

Bacterial Na(+)-translocating NADH:quinone oxidoreductase (Na(+)-NQR) uses a unique set of prosthetic redox groups-two covalently bound FMN residues, a [2Fe-2S] cluster, FAD, riboflavin and a Cys4[Fe] center-to catalyze electron transfer from NADH to ubiquinone in a reaction coupled with Na(+) translocation across the membrane. Here we used an ultra-fast microfluidic stopped-flow instrument to determine rate constants and the difference spectra for the six consecutive reaction steps of Vibrio harveyi Na(+)-NQR reduction by NADH. The instrument, with a dead time of 0.25 ms and optical path length of 1 cm allowed collection of visible spectra in 50-μs intervals. By comparing the spectra of reaction steps with the spectra of known redox transitions of individual enzyme cofactors, we were able to identify the chemical nature of most intermediates and the sequence of electron transfer events. A previously unknown spectral transition was detected and assigned to the Cys4[Fe] center reduction. Electron transfer from the [2Fe-2S] cluster to the Cys4[Fe] center and all subsequent steps were markedly accelerated when Na(+) concentration was increased from 20 μM to 25 mM, suggesting coupling of the former step with tight Na(+) binding to or occlusion by the enzyme. An alternating access mechanism was proposed to explain electron transfer between subunits NqrF and NqrC. According to the proposed mechanism, the Cys4[Fe] center is alternatively exposed to either side of the membrane, allowing the [2Fe-2S] cluster of NqrF and the FMN residue of NqrC to alternatively approach the Cys4[Fe] center from different sides of the membrane. Copyright © 2015 Elsevier B.V. All rights reserved.

Control of Ion Selectivity in LeuT: Two Na+ Binding Sites with two different mechanisms

PubMed Central

Noskov, Sergei Y.; Roux, Benoît

2016-01-01

The x-ray structure of LeuT, a bacterial homologue of Na+/Cl−-dependent neurotransmitter transporter, provides a great opportunity to better understand the molecular basis of monovalent cation selectivity in ion-coupled transporters. LeuT possesses two ion-binding sites, NA1 and NA2, which are highly selective for Na+. Extensive all-atom free energy molecular dynamics simulations of LeuT embedded in an explicit membrane are performed at different temperatures and various occupancy states of the binding sites to dissect the molecular mechanism of ion selectivity. The results show that the two binding sites display robust selectivity for Na+ over K+ or Li+, the competing ions of most similar radii. Of particular interest, the mechanism primarily responsible for selectivity for each of the two binding sites appears to be different. In site NA1, selectivity for Na+ over K+ arises predominantly from the strong electrostatic field arising from the negatively charged carboxylate group of the leucine substrate coordinating the ion directly. In site NA2, which comprises only neutral ligands, selectivity for Na+ is enforced by the local structural restraints arising from the hydrogen-bonding network and the covalent connectivity of the poly-peptide chain surrounding the ion according to a snug-fit mechanism. PMID:18280500

Increased apical Na+ permeability in cystic fibrosis is supported by a quantitative model of epithelial ion transport

PubMed Central

O’Donoghue, Donal L; Dua, Vivek; Moss, Guy W J; Vergani, Paola

2013-01-01

Cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, which encodes an anion channel. In the human lung CFTR loss causes abnormal ion transport across airway epithelial cells. As a result CF individuals produce thick mucus, suffer persistent bacterial infections and have a much reduced life expectancy. Trans-epithelial potential difference (Vt) measurements are routinely carried out on nasal epithelia of CF patients in the clinic. CF epithelia exhibit a hyperpolarised basal Vt and a larger Vt change in response to amiloride (a blocker of the epithelial Na+ channel, ENaC). Are these altered bioelectric properties solely a result of electrical coupling between the ENaC and CFTR currents, or are they due to an increased ENaC permeability associated with CFTR loss? To examine these issues we have developed a quantitative mathematical model of human nasal epithelial ion transport. We find that while the loss of CFTR permeability hyperpolarises Vt and also increases amiloride-sensitive Vt, these effects are too small to account for the magnitude of change observed in CF epithelia. Instead, a parallel increase in ENaC permeability is required to adequately fit observed experimental data. Our study provides quantitative predictions for the complex relationships between ionic permeabilities and nasal Vt, giving insights into the physiology of CF disease that have important implications for CF therapy. PMID:23732645

A constitutive pan-hexose permease for the Plasmodium life cycle and transgenic models for screening of antimalarial sugar analogs.

PubMed

Blume, Martin; Hliscs, Marion; Rodriguez-Contreras, Dayana; Sanchez, Marco; Landfear, Scott; Lucius, Richard; Matuschewski, Kai; Gupta, Nishith

2011-04-01

Glucose is considered essential for erythrocytic stages of the malaria parasite, Plasmodium falciparum. Importance of sugar and its permease for hepatic and sexual stages of Plasmodium, however, remains elusive. Moreover, increasing global resistance to current antimalarials necessitates the search for novel drugs. Here, we reveal that hexose transporter 1 (HT1) of Plasmodium berghei can transport glucose (K(m)~87 μM), mannose (K(i)~93 μM), fructose (K(i)~0.54 mM), and galactose (K(i)~5 mM) in Leishmania mexicana mutant and Xenopus laevis; and, therefore, is functionally equivalent to HT1 of P. falciparum (Glc, K(m)~175 μM; Man, K(i)~276 μM; Fru, K(i)~1.25 mM; Gal, K(i)~5.86 mM). Notably, a glucose analog, C3361, attenuated hepatic (IC(50)~15 μM) and ookinete development of P. berghei. The PbHT1 could be ablated during intraerythrocytic stages only by concurrent complementation with PbHT1-HA or PfHT1. Together; these results signify that PbHT1 and glucose are required for the entire life cycle of P. berghei. Accordingly, PbHT1 is expressed in the plasma membrane during all parasite stages. To permit a high-throughput screening of PfHT1 inhibitors and their subsequent in vivo assessment, we have generated Saccharomyces cerevisiae mutant expressing codon-optimized PfHT1, and a PfHT1-dependent Δpbht1 parasite strain. This work provides a platform to facilitate the development of drugs against malaria, and it suggests a disease-control aspect by reducing parasite transmission.

Cellular cation transport studied by 6/7Li and 23Na NMR in a porous Mo132 Keplerate type nano-capsule as model system.

PubMed

Rehder, Dieter; Haupt, Erhard T K; Müller, Achim

2008-01-01

Li+ ions can interplay with other cations intrinsically present in the intra- and extra-cellular space (i.e. Na+, K+, Mg2+ and Ca2+) have therapeutic effects (e.g. in the treatment of bipolar disorder) or toxic effects (at higher doses), likely because Li+ interferes with the intra-/extra-cellular concentration gradients of the mentioned physiologically relevant cations. The cellular transmembrane transport can be modelled by molybdenum-oxide-based Keplerates, i.e. nano-sized porous capsules containing 132 Mo centres, monitored through 6/7Li as well as 23Na NMR spectroscopy. The effects on the transport of Li+ cations through the 'ion channels' of these model cells, caused by variations in water amount, temperature, and by the addition of organic cationic 'plugs' and the shift reagent [Dy(PPP)2](7-) are reported. In the investigated solvent systems, water acts as a transport mediator for Li+. Likewise, the counter-transport (Li+/Na+, Li+/K+, Li+/Cs+ and Li+/Ca2+) has been investigated by 7Li NMR and, in the case of Li+/Na+ exchange, by 23Na NMR, and it has been shown that most (in the case of Na+ and K+, all (Ca2+) or almost none (Cs+) of the Li cations is extruded from the internal sites of the artificial cell to the extra-cellular medium, while Na+, K+ and Ca2+ are partially incorporated.

Transport properties of coupled quantum dots in the presence of phonons

NASA Astrophysics Data System (ADS)

Martins, G.; Al-Hassanieh, K.

2005-03-01

Here is presented the numerical study of the effect of Holstein phonons in the transport properties of two coupled quantum dots (QDs) in the Kondo regime. For the QDs we use the Anderson impurity model and each QD is coupled to a different Holstein mode. At T=0, in the absence of phonons, and with 1 electron per dot, the usual splitting of the Kondo resonance is observed.^1 When the QDs are coupled to the phonons, there is a reduction of the effective Coulomb repulsion, which is explained through a canonical transformation. In addition, the conductance at the electron-hole symmetric gate potential is not affected by the phonons. This is caused by the modulation of the coupling factors.^2 The difference between the effects of phonons in lithographic QDs and in molecular conductors is also discussed. 1- C.A. Büsser et al, Phys. Rev. B 62, 9907 (2000). 2- K.A. Al-Hassanieh, C.A. Büsser, G.B. Martins, Adriana Moreo and Elbio Dagotto (preprint)

The bacterial dicarboxylate transporter, VcINDY, uses a two-domain elevator-type mechanism

PubMed Central

Mulligan, Christopher; Fenollar-Ferrer, Cristina; Fitzgerald, Gabriel A.; Vergara-Jaque, Ariela; Kaufmann, Desirée; Li, Yan; Forrest, Lucy R.; Mindell, Joseph A.

2016-01-01

Secondary transporters use alternating access mechanisms to couple uphill substrate movement to downhill ion flux. Most known transporters utilize a “rocking bundle” motion, where the protein moves around an immobile substrate binding site. However, the glutamate transporter homolog, GltPh, translocates its substrate binding site vertically across the membrane, an “elevator” mechanism. Here, we used the “repeat swap” approach to computationally predict the outward-facing state of the Na+/succinate transporter VcINDY, from Vibrio cholerae. Our model predicts a substantial “elevator”-like movement of vcINDY’s substrate binding site, with a vertical translation of ~15 Å and a rotation of ~43°; multiple disulfide crosslinks which completely inhibit transport provide experimental confirmation and demonstrate that such movement is essential. In contrast, crosslinks across the VcINDY dimer interface preserve transport, revealing an absence of large scale coupling between protomers. PMID:26828963

Asymptotic Analysis of Time-Dependent Neutron Transport Coupled with Isotopic Depletion and Radioactive Decay

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brantley, P S

2006-09-27

We describe an asymptotic analysis of the coupled nonlinear system of equations describing time-dependent three-dimensional monoenergetic neutron transport and isotopic depletion and radioactive decay. The classic asymptotic diffusion scaling of Larsen and Keller [1], along with a consistent small scaling of the terms describing the radioactive decay of isotopes, is applied to this coupled nonlinear system of equations in a medium of specified initial isotopic composition. The analysis demonstrates that to leading order the neutron transport equation limits to the standard time-dependent neutron diffusion equation with macroscopic cross sections whose number densities are determined by the standard system of ordinarymore » differential equations, the so-called Bateman equations, describing the temporal evolution of the nuclide number densities.« less

Exact Thermal Transport Properties of Gray-Arsenic using Electon-Phonon Coupling

NASA Astrophysics Data System (ADS)

Kang, Seoung-Hun; Kwon, Young-Kyun

Using various theoretical methods, we investigate the thermoelectric property of gray arsenic. Thermoelectric devices that utilize the Seebeck effect convert heat flow into electrical energy. The conversion efficiency of such a device is determined by its figure of merit or ZT value, which is related to various transport coefficients, such as Seebeck coefficient and the ratio of its electrical conductivity to its thermal counterpart for given temperature. To calculate various transport coefficients and thus the ZT values of gray arsenic, we apply the Boltzmann transport theory to its electronic and phononic structures obtained by density functional theory and density functional perturbation theory together with maximally locallized Wannier functions. During this procedure, we evaluate its relaxation time accurately by explicitly considering electron-phonon coupling. Our result reveals that gray arsenic may be used for a good p-type thermoelectric devices.

High-resolution molecular-beam spectroscopy of NaCN and Na 13CN

NASA Astrophysics Data System (ADS)

van Vaals, J. J.; Meerts, W. Leo; Dymanus, A.

The sodium cyanide molecule was studied by molecular-beam electric-resonance spectroscopy in the microwave region. We used the seeded-beam technique to produce a supersonic beam with strong translational, rotational and vibrational cooling. In the frequency range 9.5-40 GHz we observed and identified for NaCN 186 and for Na 13CN 107 hyperfine transitions in 20 and 16 rotational transitions, respectively, all in the ground vibrational state. The rotational, the five quartic and three sextic centrifugal distortion constants of NaCN are: A″ = 57921.954(7) MHz; B″ = 8369.312(2) MHz, C″ = 7272.712(2) MHz. All quadrupole and several spin-rotation coupling constants for the hyperfine interaction were evaluated. The quadrupole coupling constants (in MHz) for NaCN are: eQq12(Na) = -5.344(5), eQq12 = 2.397(7). eQq12(N) = 2.148(4), eQq12(N) = -4.142(5). From these constants and those of Na 13CN we have determined the principal components of the quadrupole coupling tensor for potassium and nitrogen. The structure of sodium cyanide evaluated from the rotational constants of NaCN and Na 13CN was found to be T shaped, similar to the structure of KCN but completely different from the linear isocyanide configuration of LiNC. The effective structural parameters for sodium cyanide in the ground vibrational state are: rCN = 1.170(4) Å, rNaC = 2.379(15) Å, rN12N = 2.233(15) Å, in gratifying agreement with ab initio calculations. Both the geometrical structure and the hyperfine coupling justify the conclusion that the CN group in gaseous sodium cyanide approximately can be considered as a free CN - ion.

Validation of a coupled core-transport, pedestal-structure, current-profile and equilibrium model

NASA Astrophysics Data System (ADS)

Meneghini, O.

2015-11-01

The first workflow capable of predicting the self-consistent solution to the coupled core-transport, pedestal structure, and equilibrium problems from first-principles and its experimental tests are presented. Validation with DIII-D discharges in high confinement regimes shows that the workflow is capable of robustly predicting the kinetic profiles from on axis to the separatrix and matching the experimental measurements to within their uncertainty, with no prior knowledge of the pedestal height nor of any measurement of the temperature or pressure. Self-consistent coupling has proven to be essential to match the experimental results, and capture the non-linear physics that governs the core and pedestal solutions. In particular, clear stabilization of the pedestal peeling ballooning instabilities by the global Shafranov shift and destabilization by additional edge bootstrap current, and subsequent effect on the core plasma profiles, have been clearly observed and documented. In our model, self-consistency is achieved by iterating between the TGYRO core transport solver (with NEO and TGLF for neoclassical and turbulent flux), and the pedestal structure predicted by the EPED model. A self-consistent equilibrium is calculated by EFIT, while the ONETWO transport package evolves the current profile and calculates the particle and energy sources. The capabilities of such workflow are shown to be critical for the design of future experiments such as ITER and FNSF, which operate in a regime where the equilibrium, the pedestal, and the core transport problems are strongly coupled, and for which none of these quantities can be assumed to be known. Self-consistent core-pedestal predictions for ITER, as well as initial optimizations, will be presented. Supported by the US Department of Energy under DE-FC02-04ER54698, DE-SC0012652.

Interfacial thermal transport with strong system-bath coupling: A phonon delocalization effect

NASA Astrophysics Data System (ADS)

He, Dahai; Thingna, Juzar; Cao, Jianshu

2018-05-01

We study the effect of system-bath coupling strength on quantum thermal transport through the interface of two weakly coupled anharmonic molecular chains by using a quantum self-consistent phonon approach. The approach inherently assumes that the two segments (anharmonic molecular chains) are approximately in local thermal equilibrium with respect to the baths that they are connected to and transforms the strongly anharmonic system into an effective harmonic one with a temperature-dependent transmission. Despite the approximations, the approach is ideal for our setup, wherein the weak interfacial coupling guarantees an approximate local thermal equilibrium of each segment and short chain length (less than the phonon mean-free path) ensues from the effective harmonic approximation. Remarkably, the heat current shows a resonant to bi-resonant transition due to the variations in the interfacial coupling and temperature, which is attributed to the delocalization of phonon modes. Delocalization occurs only in the strong system-bath coupling regime and we utilize it to model a thermal rectifier whose ratio can be nonmonotonically tuned not only with the intrinsic system parameters but also with the external temperature.

Review. The mammalian proton-coupled peptide cotransporter PepT1: sitting on the transporter-channel fence?

PubMed

Meredith, David

2009-01-27

The proton-coupled di- and tripeptide transporter PepT1 (SLC15a1) is the major route by which dietary nitrogen is taken up from the small intestine, as well as being the route of entry for important therapeutic (pro)drugs such as the beta-lactam antibiotics, angiotensin-converting enzyme inhibitors and antiviral and anti-cancer agents. PepT1 is a member of the major facilitator superfamily of 12 transmembrane domain transporter proteins. Expression studies in Xenopus laevis on rabbit PepT1 that had undergone site-directed mutagenesis of a conserved arginine residue (arginine282 in transmembrane domain 7) to a glutamate revealed that this residue played a role in the coupling of proton and peptide transport and prevented the movement of non-coupled ions during the transporter cycle. Mutations of arginine282 to other non-positive residues did not uncouple proton-peptide cotransport, but did allow additional ion movements when substrate was added. By contrast, mutations to positive residues appeared to function the same as wild-type. These findings are discussed in relation to the functional role that arginine282 may play in the way PepT1 operates, together with structural information from the homology model of PepT1 based on the Escherichia coli lactose permease crystal structure.

Transport through an impurity tunnel coupled to a Si/SiGe quantum dot

DOE PAGES

Foote, Ryan H.; Ward, Daniel R.; Prance, J. R.; ...

2015-09-11

Achieving controllable coupling of dopants in silicon is crucial for operating donor-based qubit devices, but it is difficult because of the small size of donor-bound electron wavefunctions. Here in this paper, we report the characterization of a quantum dot coupled to a localized electronic state and present evidence of controllable coupling between the quantum dot and the localized state. A set of measurements of transport through the device enable the determination that the most likely location of the localized state is consistent with a location in the quantum well near the edge of the quantum dot. Finally, our results aremore » consistent with a gate-voltage controllable tunnel coupling, which is an important building block for hybrid donor and gate-defined quantum dot devices.« less

Na+/H+ exchanger 3 inhibitor diminishes hepcidin-enhanced duodenal calcium transport in hemizygous β-globin knockout thalassemic mice.

PubMed

Charoenphandhu, Narattaphol; Kraidith, Kamonshanok; Lertsuwan, Kornkamon; Sripong, Chanakarn; Suntornsaratoon, Panan; Svasti, Saovaros; Krishnamra, Nateetip; Wongdee, Kannikar

2017-03-01

Recent investigation has shown that the liver-derived iron-regulating hormone, hepcidin, can potentiate intestinal calcium absorption in hemizygous β-globin knockout thalassemic (BKO) mice. Since the upregulation of Fe 2+ and H + cotransporter, divalent metal transporter (DMT)-1, has been shown to correlate with thalassemia-induced intestinal calcium absorption impairment, the inhibition of the apical Na + /H + exchanger (NHE)-3 that is essential for cytoplasmic pH regulation and transepithelial sodium absorption was hypothesized to negatively affect hepcidin action. Herein, the positive effect of hepcidin on the duodenal calcium transport was evaluated using Ussing chamber technique. The results showed that BKO mice had lower absorptive surface area and duodenal calcium transport than wild-type mice. Besides, paracellular transport of zinc in BKO mice was compromised. Hepcidin administration completely restored calcium transport. Since this hepcidin action was totally abolished by inhibitors of the basolateral calcium transporters, Na + /Ca 2+ exchanger (NCX1) and plasma membrane Ca 2+ -ATPase (PMCA 1b ), the enhanced calcium flux potentially occurred through the transcellular pathway rather than paracellular pathway. Interestingly, the selective NHE3 inhibitor, 100 nM tenapanor, markedly inhibited hepcidin-enhanced calcium transport. Accordingly, hepcidin is one of the promising therapeutic agents for calcium malabsorption in β-thalassemia. It mainly stimulates the transcellular calcium transport across the duodenal epithelium in an NHE3-dependent manner.

Spin-orbit coupling, electron transport and pairing instabilities in two-dimensional square structures

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kocharian, Armen N.; Fernando, Gayanath W.; Fang, Kun

Rashba spin-orbit effects and electron correlations in the two-dimensional cylindrical lattices of square geometries are assessed using mesoscopic two-, three- and four-leg ladder structures. Here the electron transport properties are systematically calculated by including the spin-orbit coupling in tight binding and Hubbard models threaded by a magnetic flux. These results highlight important aspects of possible symmetry breaking mechanisms in square ladder geometries driven by the combined effect of a magnetic gauge field spin-orbit interaction and temperature. The observed persistent current, spin and charge polarizations in the presence of spin-orbit coupling are driven by separation of electron and hole charges andmore » opposite spins in real-space. The modeled spin-flip processes on the pairing mechanism induced by the spin-orbit coupling in assembled nanostructures (as arrays of clusters) engineered in various two-dimensional multi-leg structures provide an ideal playground for understanding spatial charge and spin density inhomogeneities leading to electron pairing and spontaneous phase separation instabilities in unconventional superconductors. Such studies also fall under the scope of current challenging problems in superconductivity and magnetism, topological insulators and spin dependent transport associated with numerous interfaces and heterostructures.« less

Design of 150W, 105-μm, 0.22NA, fiber coupled laser diode module by ZEMAX

NASA Astrophysics Data System (ADS)

Qi, Yunfei; Zhao, Pengfei; Chen, Qing; Wu, Yulong; Chen, Yongqi; Zou, Yonggang; Lin, Xuechun

2016-10-01

We represent a design of a high brightness, fiber coupled diode laser module based on 16 single emitters at 915nm. The module can produce more than 150 Watts output power from a standard fiber with core diameter of 105μm and numerical aperture (NA) of 0.22. To achieve a high power and high brightness laser beam, the spatial beam combination and polarization beam combination are used to combine output of 16 single emitters into a single beam, and then an aspheric lens is used to couple the combined beam into an optical fiber. The simulation show that the total coupling efficiency is more than 95% and the highest brightness is estimated to be 11MW/ (cm2*sr).

Quantification of Na+,K+ pumps and their transport rate in skeletal muscle: Functional significance

PubMed Central

2013-01-01

During excitation, muscle cells gain Na+ and lose K+, leading to a rise in extracellular K+ ([K+]o), depolarization, and loss of excitability. Recent studies support the idea that these events are important causes of muscle fatigue and that full use of the Na+,K+-ATPase (also known as the Na+,K+ pump) is often essential for adequate clearance of extracellular K+. As a result of their electrogenic action, Na+,K+ pumps also help reverse depolarization arising during excitation, hyperkalemia, and anoxia, or from cell damage resulting from exercise, rhabdomyolysis, or muscle diseases. The ability to evaluate Na+,K+-pump function and the capacity of the Na+,K+ pumps to fill these needs require quantification of the total content of Na+,K+ pumps in skeletal muscle. Inhibition of Na+,K+-pump activity, or a decrease in their content, reduces muscle contractility. Conversely, stimulation of the Na+,K+-pump transport rate or increasing the content of Na+,K+ pumps enhances muscle excitability and contractility. Measurements of [3H]ouabain binding to skeletal muscle in vivo or in vitro have enabled the reproducible quantification of the total content of Na+,K+ pumps in molar units in various animal species, and in both healthy people and individuals with various diseases. In contrast, measurements of 3-O-methylfluorescein phosphatase activity associated with the Na+,K+-ATPase may show inconsistent results. Measurements of Na+ and K+ fluxes in intact isolated muscles show that, after Na+ loading or intense excitation, all the Na+,K+ pumps are functional, allowing calculation of the maximum Na+,K+-pumping capacity, expressed in molar units/g muscle/min. The activity and content of Na+,K+ pumps are regulated by exercise, inactivity, K+ deficiency, fasting, age, and several hormones and pharmaceuticals. Studies on the α-subunit isoforms of the Na+,K+-ATPase have detected a relative increase in their number in response to exercise and the glucocorticoid dexamethasone but have not

The Effect of Yaw Coupling in Turning Maneuvers of Large Transport Aircraft

NASA Technical Reports Server (NTRS)

McNeill, Walter E.; Innis, Robert C.

1965-01-01

A study has been made, using a piloted moving simulator, of the effects of the yaw-coupling parameters N(sub p) and N(sub delta(sub a) on the lateral-directional handling qualities of a large transport airplane at landing-approach airspeed. It is shown that the desirable combinations of these parameters tend to be more proverse when compared with values typical of current aircraft. Results of flight tests in a large variable-stability jet transport showed trends which were similar to those of the simulator data. Areas of minor disagreement, which were traced to differences in airplane geometry, indicate that pilot consciousness of side acceleration forces can be an important factor in handling qualities of future long-nosed transport aircraft.

Coupling continuum dislocation transport with crystal plasticity for application to shock loading conditions

DOE PAGES

Luscher, Darby Jon; Mayeur, Jason Rhea; Mourad, Hashem Mohamed; ...

2015-08-05

Here, we have developed a multi-physics modeling approach that couples continuum dislocation transport, nonlinear thermoelasticity, crystal plasticity, and consistent internal stress and deformation fields to simulate the single-crystal response of materials under extreme dynamic conditions. Dislocation transport is modeled by enforcing dislocation conservation at a slip-system level through the solution of advection-diffusion equations. Nonlinear thermoelasticity provides a thermodynamically consistent equation of state to relate stress (including pressure), temperature, energy densities, and dissipation. Crystal plasticity is coupled to dislocation transport via Orowan's expression where the constitutive description makes use of recent advances in dislocation velocity theories applicable under extreme loading conditions.more » The configuration of geometrically necessary dislocation density gives rise to an internal stress field that can either inhibit or accentuate the flow of dislocations. An internal strain field associated with the internal stress field contributes to the kinematic decomposition of the overall deformation. The paper describes each theoretical component of the framework, key aspects of the constitutive theory, and some details of a one-dimensional implementation. Results from single-crystal copper plate impact simulations are discussed in order to highlight the role of dislocation transport and pile-up in shock loading regimes. The main conclusions of the paper reinforce the utility of the modeling approach to shock problems.« less

NUTRIENT SOUIRCES, TRANSPORT, AND FATE IN COUPLED WATERSHED-ESTUARINE SYSTEMS OF COASTAL ALABAMA

EPA Science Inventory

The processes regulating sources, transport, and fate of nutrients were studied in 3 coupled watershed-estuarine systems that varied mainly by differences in the dominant land use-land cover (LULC), i.e. Weeks Bay -- agriculture, Dog River -- urban, and Fowl River -- forest. Mea...

Quantum transport properties of carbon nanotube field-effect transistors with electron-phonon coupling

NASA Astrophysics Data System (ADS)

Ishii, Hiroyuki; Kobayashi, Nobuhiko; Hirose, Kenji

2007-11-01

We investigated the electron-phonon coupling effects on the electronic transport properties of metallic (5,5)- and semiconducting (10,0)-carbon nanotube devices. We calculated the conductance and mobility of the carbon nanotubes with micron-order lengths at room temperature, using the time-dependent wave-packet approach based on the Kubo-Greenwood formula within a tight-binding approximation. We investigated the scattering effects of both longitudinal acoustic and optical phonon modes on the transport properties. The electron-optical phonon coupling decreases the conductance around the Fermi energy for the metallic carbon nanotubes, while the conductance of semiconductor nanotubes is decreased around the band edges by the acoustic phonons. Furthermore, we studied the Schottky-barrier effects on the mobility of the semiconducting carbon nanotube field-effect transistors for various gate voltages. We clarified how the electron mobilities of the devices are changed by the acoustic phonon.

Modeling coupled nanoparticle aggregation and transport in porous media: A Lagrangian approach

NASA Astrophysics Data System (ADS)

Taghavy, Amir; Pennell, Kurt D.; Abriola, Linda M.

2015-01-01

Changes in nanoparticle size and shape due to particle-particle interactions (i.e., aggregation or agglomeration) may significantly alter particle mobility and retention in porous media. To date, however, few modeling studies have considered the coupling of transport and particle aggregation processes. The majority of particle transport models employ an Eulerian modeling framework and are, consequently, limited in the types of collisions and aggregate sizes that can be considered. In this work, a more general Lagrangian modeling framework is developed and implemented to explore coupled nanoparticle aggregation and transport processes. The model was verified through comparison of model simulations to published results of an experimental and Eulerian modeling study (Raychoudhury et al., 2012) of carboxymethyl cellulose (CMC)-modified nano-sized zero-valent iron particle (nZVI) transport and retention in water-saturated sand columns. A model sensitivity analysis reveals the influence of influent particle concentration (ca. 70 to 700 mg/L), primary particle size (10-100 nm) and pore water velocity (ca. 1-6 m/day) on particle-particle, and, consequently, particle-collector interactions. Model simulations demonstrate that, when environmental conditions promote particle-particle interactions, neglecting aggregation effects can lead to under- or over-estimation of nanoparticle mobility. Results also suggest that the extent to which higher order particle-particle collisions influence aggregation kinetics will increase with the fraction of primary particles. This work demonstrates the potential importance of time-dependent aggregation processes on nanoparticle mobility and provides a numerical model capable of capturing/describing these interactions in water-saturated porous media.

Modeling coupled nanoparticle aggregation and transport in porous media: a Lagrangian approach.

PubMed

Taghavy, Amir; Pennell, Kurt D; Abriola, Linda M

2015-01-01

Changes in nanoparticle size and shape due to particle-particle interactions (i.e., aggregation or agglomeration) may significantly alter particle mobility and retention in porous media. To date, however, few modeling studies have considered the coupling of transport and particle aggregation processes. The majority of particle transport models employ an Eulerian modeling framework and are, consequently, limited in the types of collisions and aggregate sizes that can be considered. In this work, a more general Lagrangian modeling framework is developed and implemented to explore coupled nanoparticle aggregation and transport processes. The model was verified through comparison of model simulations to published results of an experimental and Eulerian modeling study (Raychoudhury et al., 2012) of carboxymethyl cellulose (CMC)-modified nano-sized zero-valent iron particle (nZVI) transport and retention in water-saturated sand columns. A model sensitivity analysis reveals the influence of influent particle concentration (ca. 70 to 700 mg/L), primary particle size (10-100 nm) and pore water velocity (ca. 1-6 m/day) on particle-particle, and, consequently, particle-collector interactions. Model simulations demonstrate that, when environmental conditions promote particle-particle interactions, neglecting aggregation effects can lead to under- or over-estimation of nanoparticle mobility. Results also suggest that the extent to which higher order particle-particle collisions influence aggregation kinetics will increase with the fraction of primary particles. This work demonstrates the potential importance of time-dependent aggregation processes on nanoparticle mobility and provides a numerical model capable of capturing/describing these interactions in water-saturated porous media. Copyright © 2014 Elsevier B.V. All rights reserved.

Targeting the AtCWIN1 Gene to Explore the Role of Invertases in Sucrose Transport in Roots and during Botrytis cinerea Infection

PubMed Central

Veillet, Florian; Gaillard, Cécile; Coutos-Thévenot, Pierre; La Camera, Sylvain

2016-01-01

Cell wall invertases (CWIN) cleave sucrose into glucose and fructose in the apoplast. CWINs are key regulators of carbon partitioning and source/sink relationships during growth, development and under biotic stresses. In this report, we monitored the expression/activity of Arabidopsis cell wall invertases in organs behaving as source, sink, or subjected to a source/sink transition after infection with the necrotrophic fungus Botrytis cinerea. We showed that organs with different source/sink status displayed differential CWIN activities, depending on carbohydrate needs or availabilities in the surrounding environment, through a transcriptional and posttranslational regulation. Loss-of-function mutation of the Arabidopsis cell wall invertase 1 gene, AtCWIN1, showed that the corresponding protein was the main contributor to the apoplastic sucrose cleaving activity in both leaves and roots. The CWIN-deficient mutant cwin1-1 exhibited a reduced capacity to actively take up external sucrose in roots, indicating that this process is mainly dependent on the sucrolytic activity of AtCWIN1. Using T-DNA and CRISPR/Cas9 mutants impaired in hexose transport, we demonstrated that external sucrose is actively absorbed in the form of hexoses by a sugar/H+ symport system involving the coordinated activity of AtCWIN1 with several Sugar Transporter Proteins (STP) of the plasma membrane, i.e., STP1 and STP13. Part of external sucrose was imported without apoplastic cleavage into cwin1-1 seedling roots, highlighting an alternative AtCWIN1-independent pathway for the assimilation of external sucrose. Accordingly, we showed that several genes encoding sucrose transporters of the plasma membrane were expressed. We also detected transcript accumulation of vacuolar invertase (VIN)-encoding genes and high VIN activities. Upon infection, AtCWIN1 was responsible for all the Botrytis-induced apoplastic invertase activity. We detected a transcriptional activation of several AtSUC and AtVIN genes

Localization and Function of the Membrane-bound Riboflavin in the Na+-translocating NADH:Quinone Oxidoreductase (Na+-NQR) from Vibrio cholerae*

PubMed Central

Casutt, Marco S.; Huber, Tamara; Brunisholz, René; Tao, Minli; Fritz, Günter; Steuber, Julia

2010-01-01

The sodium ion-translocating NADH:quinone oxidoreductase (Na+-NQR) from the human pathogen Vibrio cholerae is a respiratory membrane protein complex that couples the oxidation of NADH to the transport of Na+ across the bacterial membrane. The Na+-NQR comprises the six subunits NqrABCDEF, but the stoichiometry and arrangement of these subunits are unknown. Redox-active cofactors are FAD and a 2Fe-2S cluster on NqrF, covalently attached FMNs on NqrB and NqrC, and riboflavin and ubiquinone-8 with unknown localization in the complex. By analyzing the cofactor content and NADH oxidation activity of subcomplexes of the Na+-NQR lacking individual subunits, the riboflavin cofactor was unequivocally assigned to the membrane-bound NqrB subunit. Quantitative analysis of the N-terminal amino acids of the holo-complex revealed that NqrB is present in a single copy in the holo-complex. It is concluded that the hydrophobic NqrB harbors one riboflavin in addition to its covalently attached FMN. The catalytic role of two flavins in subunit NqrB during the reduction of ubiquinone to ubiquinol by the Na+-NQR is discussed. PMID:20558724

Kinetic contribution to extracellular Na+/K+ selectivity in the Na+/K+ pump.

PubMed

Vleeskens, Elizabeth; Clarke, Ronald J

2018-05-01

The sodium potassium pump (Na + ,K + -ATPase) shows a high selectivity for K + over Na + binding from the extracellular medium. To understand the K + selectivity in the presence of a high concentration of competing Na + ions requires consideration of more than just ion binding affinities. Here, equilibrium-based calculations of the extracellular occupation of the Na + ,K + -ATPase transport sites by Na + and K + are compared to fluxes through Na + and K + transport pathways. The results show that, under physiological conditions, there is a 332-fold selectivity for pumping of K + from the extracellular medium into the cytoplasm relative to Na + , whereas equilibrium calculations alone predict only a 7.5-fold selectivity for K + . Thus, kinetic effects make a major contribution to the determination of extracellular K + selectivity.

Inhibition of epithelial Na sup + transport by atriopeptin, protein kinase c, and pertussis toxin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mohrmann, M.; Cantiello, H.F.; Ausiello, D.A.

1987-08-01

The authors have recently shown the selective inhibition of an amiloride-sensitive, conductive pathway for Na{sup +} by atrial natriuretic peptide and 8-bromoguanosine 3{prime},5{prime}-cyclic monophosphate (8-BrcGMP) in the renal epithelial cell line, LLC-PK{sub i}. Using {sup 22}Na{sup +} fluxes, they further investigated the modulation of Na{sup +} transport by atrial natriuretic peptide and by agents that increase cGMP production, activate protein kinase c, or modulate guanine nucleotide regulatory protein function. Sodium nitroprusside increases intracellular cGMP concentrations without affecting cAMP concentrations and completely inhibits amiloride-sensitive Na{sup +} uptake in a time- and concentration-dependent manner. Oleoyl 2-acetylglycerol and phorbol 12-myristate 13-acetate, activators ofmore » protein kinase c, inhibit Na{sup +} uptake by 93 {plus minus} 13 and 51 {plus minus} 10%, respectively. Prolonged incubation with phorbol ester results in the downregulation of protein kinase c activity and reduces the inhibitory effect of atrial natriuretic peptide, suggesting that the action of this peptide involves stimulation of protein kinase c. Pertussis toxin, which induces the ADP-ribosylation of a 41-kDa guanine nucleotide regulatory protein in LLC-PK{sub i} cells, inhibits {sup 22}Na{sup +} influx to the same extent as amiloride. Thus, increasing cGMP, activating protein kinase c, and ADP-ribosylating a guanine nucleotide regulatory protein all inhibit Na{sup +} uptake. These events may be sequentially involved in the action of atrial natriuretic peptide.« less

Insights from the Fungus Fusarium oxysporum Point to High Affinity Glucose Transporters as Targets for Enhancing Ethanol Production from Lignocellulose

PubMed Central

Ali, Shahin S.; Nugent, Brian; Mullins, Ewen; Doohan, Fiona M.

2013-01-01

Ethanol is the most-widely used biofuel in the world today. Lignocellulosic plant biomass derived from agricultural residue can be converted to ethanol via microbial bioprocessing. Fungi such as Fusarium oxysporum can simultaneously saccharify straw to sugars and ferment sugars to ethanol. But there are many bottlenecks that need to be overcome to increase the efficacy of microbial production of ethanol from straw, not least enhancement of the rate of fermentation of both hexose and pentose sugars. This research tested the hypothesis that the rate of sugar uptake by F. oxysporum would enhance the ethanol yields from lignocellulosic straw and that high affinity glucose transporters can enhance ethanol yields from this substrate. We characterized a novel hexose transporter (Hxt) from this fungus. The F. oxysporum Hxt represents a novel transporter with homology to yeast glucose signaling/transporter proteins Rgt2 and Snf3, but it lacks their C-terminal domain which is necessary for glucose signalling. Its expression level decreased with increasing glucose concentration in the medium and in a glucose uptake study the Km(glucose) was 0.9 mM, which indicated that the protein is a high affinity glucose transporter. Post-translational gene silencing or over expression of the Hxt in F. oxysporum directly affected the glucose and xylose transport capacity and ethanol yielded by F. oxysporum from straw, glucose and xylose. Thus we conclude that this Hxt has the capacity to transport both C5 and C6 sugars and to enhance ethanol yields from lignocellulosic material. This study has confirmed that high affinity glucose transporters are ideal candidates for improving ethanol yields from lignocellulose because their activity and level of expression is high in low glucose concentrations, which is very common during the process of consolidated processing. PMID:23382943

H(+)/solute-induced intracellular acidification leads to selective activation of apical Na(+)/H(+) exchange in human intestinal epithelial cells.

PubMed

Thwaites, D T; Ford, D; Glanville, M; Simmons, N L

1999-09-01

The intestinal absorption of many nutrients and drug molecules is mediated by ion-driven transport mechanisms in the intestinal enterocyte plasma membrane. Clearly, the establishment and maintenance of the driving forces - transepithelial ion gradients - are vital for maximum nutrient absorption. The purpose of this study was to determine the nature of intracellular pH (pH(i)) regulation in response to H(+)-coupled transport at the apical membrane of human intestinal epithelial Caco-2 cells. Using isoform-specific primers, mRNA transcripts of the Na(+)/H(+) exchangers NHE1, NHE2, and NHE3 were detected by RT-PCR, and identities were confirmed by sequencing. The functional profile of Na(+)/H(+) exchange was determined by a combination of pH(i), (22)Na(+) influx, and EIPA inhibition experiments. Functional NHE1 and NHE3 activities were identified at the basolateral and apical membranes, respectively. H(+)/solute-induced acidification (using glycylsarcosine or beta-alanine) led to Na(+)-dependent, EIPA-inhibitable pH(i) recovery or EIPA-inhibitable (22)Na(+) influx at the apical membrane only. Selective activation of apical (but not basolateral) Na(+)/H(+) exchange by H(+)/solute cotransport demonstrates that coordinated activity of H(+)/solute symport with apical Na(+)/H(+) exchange optimizes the efficient absorption of nutrients and Na(+), while maintaining pH(i) and the ion gradients involved in driving transport.

Tight coupling of Na+/K+-ATPase with glycolysis demonstrated in permeabilized rat cardiomyocytes.

PubMed

Sepp, Mervi; Sokolova, Niina; Jugai, Svetlana; Mandel, Merle; Peterson, Pearu; Vendelin, Marko

2014-01-01

The effective integrated organization of processes in cardiac cells is achieved, in part, by the functional compartmentation of energy transfer processes. Earlier, using permeabilized cardiomyocytes, we demonstrated the existence of tight coupling between some of cardiomyocyte ATPases and glycolysis in rat. In this work, we studied contribution of two membrane ATPases and whether they are coupled to glycolysis--sarcoplasmic reticulum Ca2+ ATPase (SERCA) and plasmalemma Na+/K+-ATPase (NKA). While SERCA activity was minor in this preparation in the absence of calcium, major role of NKA was revealed accounting to ∼30% of the total ATPase activity which demonstrates that permeabilized cell preparation can be used to study this pump. To elucidate the contribution of NKA in the pool of ATPases, a series of kinetic measurements was performed in cells where NKA had been inhibited by 2 mM ouabain. In these cells, we recorded: ADP- and ATP-kinetics of respiration, competition for ADP between mitochondria and pyruvate kinase (PK), ADP-kinetics of endogenous PK, and ATP-kinetics of total ATPases. The experimental data was analyzed using a series of mathematical models with varying compartmentation levels. The results show that NKA is tightly coupled to glycolysis with undetectable flux of ATP between mitochondria and NKA. Such tight coupling of NKA to PK is in line with its increased importance in the pathological states of the heart when the substrate preference shifts to glucose.

AtSWEET4, a hexose facilitator, mediates sugar transport to axial sinks and affects plant development

PubMed Central

Liu, Xiaozhu; Zhang, Yan; Yang, Chao; Tian, Zhihong; Li, Jianxiong

2016-01-01

Plants transport photoassimilates from source organs to sink tissues through the phloem translocation pathway. In the transport phloem, sugars that escape from the sieve tubes are released into the apoplasmic space between the sieve element/companion cell complex (SE/CC) and phloem parenchyma cells (PPCs) during the process of long-distance transport. The competition for sugar acquisition between SE/CC and adjoining PPCs is mediated by plasma membrane translocators. YFP-tagged AtSWEET4 protein is localized in the plasma membrane, and PromoterAtSWEET4-GUS analysis showed that AtSWEET4 is expressed in the stele of roots and veins of leaves and flowers. Overexpression of AtSWEET4 in Arabidopsis increases plant size and accumulates more glucose and fructose. By contrast, knock-down of AtSWEET4 by RNA-interference leads to small plant size, reduction in glucose and fructose contents, chlorosis in the leaf vein network, and reduction in chlorophyll content in leaves. Yeast assays demonstrated that AtSWEET4 is able to complement both fructose and glucose transport deficiency. Transgenic plants of AtSWEET4 overexpression exhibit higher freezing tolerance and support more growth of bacterium Pseudomonas syringae pv. phaseolicola NPS3121. We conclude that AtSWEET4 plays an important role in mediating sugar transport in axial tissues during plant growth and development. PMID:27102826

Pma1 is an alkali/alkaline earth metal cation ATPase that preferentially transports Na(+) and K(+) across the Mycobacterium smegmatis plasma membrane.

PubMed

Ayala-Torres, Carlos; Novoa-Aponte, Lorena; Soto, Carlos Y

2015-07-01

Mycobacterium smegmatis Pma1 is the orthologue of M. tuberculosis P-type ATPase cation transporter CtpF, which is activated under stress conditions, such as hypoxia, starvation and response to antituberculous and toxic substances. The function of Pma1 in the mycobacterial processes across the plasma membrane has not been characterised. In this work, bioinformatic analyses revealed that Pma1 likely contains potential sites for, Na(+), K(+) and Ca(2+) binding and transport. Accordingly, RT-qPCR experiments showed that M. smegmatis pma1 transcription is stimulated by sub-lethal doses of Na(+), K(+) and Ca(2+); in addition, the ATPase activity of plasma membrane vesicles in recombinant Pma1-expressing M. smegmatis cells is stimulated by treatment with these cations. In contrast, M. smegmatis cells homologously expressing Pma1 displayed tolerance to high doses of Na(+) and K(+) but not to Ca(2+) ions. Consistently, the recombinant protein Km embedded in plasma membrane demonstrated that Ca(2+) has more affinity for Pma1 than Na(+) and K(+) ions; furthermore, the estimation of Vmax/Km suggests that Na(+) and K(+) ions are more efficiently translocated than Ca(2+). Thus, these results strongly suggest that Pma1 is a promiscuous alkali/alkaline earth cation ATPase that preferentially transports Na(+) and/or K(+) across the mycobacterial plasma membrane. Copyright © 2015 Elsevier GmbH. All rights reserved.

Energetic and dynamic analysis of transport of Na + and K + through a cyclic peptide nanotube in water and in lipid bilayers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Song, Yeonho; Lee, Ji Hye; Hwang, Hoon

Potential of mean force (PMF) profiles and position-dependent diffusion coefficients of Na + and K + are calculated to elucidate the translocation of ions through a cyclic peptide nanotube, composed of 8 × cyclo[-(D-Leu-Trp) 4-] rings, in water and in hydrated DMPC bilayers. The PMF profiles and PMF decomposition analysis for the monovalent cations show that favorable interactions of the cations with the CPN as well as the lipid bilayer and dehydration free energy penalties are two major competing factors which determine the free energy surface for ion transport through CPNs both in water and lipid bilayers, and that themore » selectivity of CPNs to cations mainly arises from favorable interaction energies of cations with CPNs and lipid bilayers that are more dominant than the dehydration penalties. Calculations of the position-dependent diffusion coefficients and dynamic friction kernels of the cations indicate that the dehydration process along with the molecular rearrangements occurring outside the channel and the coupling of the ion motions with the chain-structured water movements inside the channel lead to decrease of the diffusion coefficients far away from the channel entrance and also reduced coefficients inside the channel. Here the PMF and diffusivity profiles for Na + and K + reveal that the energetics of ion transport through the CPN are governed by global interactions of ions with all the components in the system while the diffusivity of ions through the channel is mostly determined by local interactions of ions with the confined water molecules inside the channel. Comparison of Na + and K + ion distributions based on overdamped Brownian dynamics simulations based on the PMF and diffusivity profiles with the corresponding results from molecular dynamics shows good agreement, indicating accuracy of the Bayesian inference method for determining diffusion coefficients in this application. In addition this work shows that position-dependent diffusion

Energetic and dynamic analysis of transport of Na + and K + through a cyclic peptide nanotube in water and in lipid bilayers

DOE PAGES

Song, Yeonho; Lee, Ji Hye; Hwang, Hoon; ...

2016-11-04

Potential of mean force (PMF) profiles and position-dependent diffusion coefficients of Na + and K + are calculated to elucidate the translocation of ions through a cyclic peptide nanotube, composed of 8 × cyclo[-(D-Leu-Trp) 4-] rings, in water and in hydrated DMPC bilayers. The PMF profiles and PMF decomposition analysis for the monovalent cations show that favorable interactions of the cations with the CPN as well as the lipid bilayer and dehydration free energy penalties are two major competing factors which determine the free energy surface for ion transport through CPNs both in water and lipid bilayers, and that themore » selectivity of CPNs to cations mainly arises from favorable interaction energies of cations with CPNs and lipid bilayers that are more dominant than the dehydration penalties. Calculations of the position-dependent diffusion coefficients and dynamic friction kernels of the cations indicate that the dehydration process along with the molecular rearrangements occurring outside the channel and the coupling of the ion motions with the chain-structured water movements inside the channel lead to decrease of the diffusion coefficients far away from the channel entrance and also reduced coefficients inside the channel. Here the PMF and diffusivity profiles for Na + and K + reveal that the energetics of ion transport through the CPN are governed by global interactions of ions with all the components in the system while the diffusivity of ions through the channel is mostly determined by local interactions of ions with the confined water molecules inside the channel. Comparison of Na + and K + ion distributions based on overdamped Brownian dynamics simulations based on the PMF and diffusivity profiles with the corresponding results from molecular dynamics shows good agreement, indicating accuracy of the Bayesian inference method for determining diffusion coefficients in this application. In addition this work shows that position-dependent diffusion

Critical Surface Parameters for the Oxidative Coupling of Methane over the Mn-Na-W/SiO2 Catalyst.

PubMed

Hayek, Naseem S; Lucas, Nishita S; Warwar Damouny, Christine; Gazit, Oz M

2017-11-22

The work here presents a thorough evaluation of the effect of Mn-Na-W/SiO 2 catalyst surface parameters on its performance in the oxidative coupling of methane (OCM). To do so, we used microporous dealuminated β-zeolite (Zeo), or mesoporous SBA-15 (SBA), or macroporous fumed silica (Fum) as precursors for catalyst preparation, together with Mn nitrate, Mn acetate and Na 2 WO 4 . Characterizing the catalysts by inductively coupled plasma-optical emission spectroscopy, N 2 physisorption, X-ray diffraction, high-resolution scanning electron microscopy-energy-dispersive spectroscopy, X-ray photoelectron spectroscopy, and catalytic testing enabled us to identify critical surface parameters that govern the activity and C 2 selectivity of the Mn-Na-W/SiO 2 catalyst. Although the current paradigm views the phase transition of silica to α-cristobalite as the critical step in obtaining dispersed and stable metal sites, we show that the choice of precursors is equally or even more important with respect to tailoring the right surface properties. Specifically, the SBA-based catalyst, characterized by relatively closed surface porosity, demonstrated low activity and low C 2 selectivity. By contrast, for the same composition, the Zeo-based catalyst showed an open surface pore structure, which translated up to fourfold higher activity and enhanced selectivity. By varying the overall composition of the Zeo catalysts, we show that reducing the overall W concentration reduces the size of the Na 2 WO 4 species and increases the catalytic activity linearly as much as fivefold higher than the SBA catalyst. This linear dependence correlates well to the number of interfaces between the Na 2 WO 4 and Mn 2 O 3 species. Our results combined with prior studies lead us to single out the interface between Na 2 WO 4 and Mn 2 O 3 as the most probable active site for OCM using this catalyst. Synergistic interactions between the various precursors used and the phase transition are discussed in

Reactive Transport Modeling of Induced Calcite Precipitation Reaction Fronts in Porous Media Using A Parallel, Fully Coupled, Fully Implicit Approach

NASA Astrophysics Data System (ADS)

Guo, L.; Huang, H.; Gaston, D.; Redden, G. D.; Fox, D. T.; Fujita, Y.

2010-12-01

Inducing mineral precipitation in the subsurface is one potential strategy for immobilizing trace metal and radionuclide contaminants. Generating mineral precipitates in situ can be achieved by manipulating chemical conditions, typically through injection or in situ generation of reactants. How these reactants transport, mix and react within the medium controls the spatial distribution and composition of the resulting mineral phases. Multiple processes, including fluid flow, dispersive/diffusive transport of reactants, biogeochemical reactions and changes in porosity-permeability, are tightly coupled over a number of scales. Numerical modeling can be used to investigate the nonlinear coupling effects of these processes which are quite challenging to explore experimentally. Many subsurface reactive transport simulators employ a de-coupled or operator-splitting approach where transport equations and batch chemistry reactions are solved sequentially. However, such an approach has limited applicability for biogeochemical systems with fast kinetics and strong coupling between chemical reactions and medium properties. A massively parallel, fully coupled, fully implicit Reactive Transport simulator (referred to as “RAT”) based on a parallel multi-physics object-oriented simulation framework (MOOSE) has been developed at the Idaho National Laboratory. Within this simulator, systems of transport and reaction equations can be solved simultaneously in a fully coupled, fully implicit manner using the Jacobian Free Newton-Krylov (JFNK) method with additional advanced computing capabilities such as (1) physics-based preconditioning for solution convergence acceleration, (2) massively parallel computing and scalability, and (3) adaptive mesh refinements for 2D and 3D structured and unstructured mesh. The simulator was first tested against analytical solutions, then applied to simulating induced calcium carbonate mineral precipitation in 1D columns and 2D flow cells as analogs

Investigation of hurricane Ivan using the coupled ocean-atmosphere-wave-sediment transport (COAWST) model

USGS Publications Warehouse

Zambon, Joseph B.; He, Ruoying; Warner, John C.

2014-01-01

The coupled ocean–atmosphere–wave–sediment transport (COAWST) model is used to hindcast Hurricane Ivan (2004), an extremely intense tropical cyclone (TC) translating through the Gulf of Mexico. Sensitivity experiments with increasing complexity in ocean–atmosphere–wave coupled exchange processes are performed to assess the impacts of coupling on the predictions of the atmosphere, ocean, and wave environments during the occurrence of a TC. Modest improvement in track but significant improvement in intensity are found when using the fully atmosphere–ocean-wave coupled configuration versus uncoupled (e.g., standalone atmosphere, ocean, or wave) model simulations. Surface wave fields generated in the fully coupled configuration also demonstrates good agreement with in situ buoy measurements. Coupled and uncoupled model-simulated sea surface temperature (SST) fields are compared with both in situ and remote observations. Detailed heat budget analysis reveals that the mixed layer temperature cooling in the deep ocean (on the shelf) is caused primarily by advection (equally by advection and diffusion).

Human, rat and chicken small intestinal Na+-Cl−-creatine transporter: functional, molecular characterization and localization

PubMed Central

Peral, M J; García-Delgado, M; Calonge, M L; Durán, J M; De La Horra, M C; Wallimann, T; Speer, O; Ilundáin, A A

2002-01-01

In spite of all the fascinating properties of oral creatine supplementation, the mechanism(s) mediating its intestinal absorption has(have) not been investigated. The purpose of this study was to characterize intestinal creatine transport. [14C]Creatine uptake was measured in chicken enterocytes and rat ileum, and expression of the creatine transporter CRT was examined in human, rat and chicken small intestine by reverse transcription-polymerase chain reaction, Northern blot, in situ hybridization, immunoblotting and immunohistochemistry. Results show that enterocytes accumulate creatine against its concentration gradient. This accumulation was electrogenic, Na+- and Cl−-dependent, with a probable stoichiometry of 2 Na+: 1 Cl−: 1 creatine, and inhibited by ouabain and iodoacetic acid. The kinetic study revealed a Km for creatine of 29 μm. [14C]Creatine uptake was efficiently antagonized by non-labelled creatine, guanidinopropionic acid and cyclocreatine. More distant structural analogues of creatine, such as GABA, choline, glycine, β-alanine, taurine and betaine, had no effect on intestinal creatine uptake, indicating a high substrate specificity of the creatine transporter. Consistent with these functional data, messenger RNA for CRT was detected only in the cells lining the intestinal villus. The sequences of partial clones, and of the full-length cDNA clone, isolated from human and rat small intestine were identical to previously cloned CRT cDNAs. Immunological analysis revealed that CRT protein was mainly associated with the apical membrane of the enterocytes. This study reports for the first time that mammalian and avian enterocytes express CRT along the villus, where it mediates high-affinity, Na+- and Cl−-dependent, apical creatine uptake. PMID:12433955

AMP-activated protein kinase (AMPK)–dependent and –independent pathways regulate hypoxic inhibition of transepithelial Na+ transport across human airway epithelial cells

PubMed Central

Tan, CD; Smolenski, RT; Harhun, MI; Patel, HK; Ahmed, SG; Wanisch, K; Yáñez-Muñoz, RJ; Baines, DL

2012-01-01

BACKGROUND AND PURPOSE Pulmonary transepithelial Na+ transport is reduced by hypoxia, but in the airway the regulatory mechanisms remain unclear. We investigated the role of AMPK and ROS in the hypoxic regulation of apical amiloride-sensitive Na+ channels and basolateral Na+K+ ATPase activity. EXPERIMENTAL APPROACH H441 human airway epithelial cells were used to examine the effects of hypoxia on Na+ transport, AMP : ATP ratio and AMPK activity. Lentiviral constructs were used to modify cellular AMPK abundance and activity; pharmacological agents were used to modify cellular ROS. KEY RESULTS AMPK was activated by exposure to 3% or 0.2% O2 for 60 min in cells grown in submerged culture or when fluid (0.1 mL·cm−2) was added to the apical surface of cells grown at the air–liquid interface. Only 0.2% O2 activated AMPK in cells grown at the air–liquid interface. AMPK activation was associated with elevation of cellular AMP : ATP ratio and activity of the upstream kinase LKB1. Hypoxia inhibited basolateral ouabain-sensitive Isc (Iouabain) and apical amiloride-sensitive Na+ conductance (GNa+). Modification of AMPK activity prevented the effect of hypoxia on Iouabain (Na+K+ ATPase) but not apical GNa+. Scavenging of superoxide and inhibition of NADPH oxidase prevented the effect of hypoxia on apical GNa+ (epithelial Na+ channels). CONCLUSIONS AND IMPLICATIONS Hypoxia activates AMPK-dependent and -independent pathways in airway epithelial cells. Importantly, these pathways differentially regulate apical Na+ channels and basolateral Na+K+ ATPase activity to decrease transepithelial Na+ transport. Luminal fluid potentiated the effect of hypoxia and activated AMPK, which could have important consequences in lung disease conditions. PMID:22509822

Improvement in reduced-mode (REM) diodes enable 315 W from 105-μm 0.15-NA fiber-coupled modules

NASA Astrophysics Data System (ADS)

Kanskar, M.; Bao, L.; Chen, Z.; Dawson, D.; DeVito, M.; Dong, W.; Grimshaw, M.; Guan, X.; Hemenway, M.; Martinsen, R.; Urbanek, W.; Zhang, S.

2018-02-01

High-power, high-brightness diode lasers have been pursued for many applications including fiber laser pumping, materials processing, solid-state laser pumping, and consumer electronics manufacturing. In particular, 915 nm - and 976 nm diodes are of interest as diode pumps for the kilowatt CW fiber lasers. As a result, there have been many technical thrusts for driving the diode lasers to have both high power and high brightness to achieve high-performance and reduced manufacturing costs. This paper presents our continued progress in the development of high brightness fiber-coupled product platform, nLIGHT element®. In the past decade, the power coupled into a single 105 μm and 0.15 NA fiber has increased by over a factor of ten through improved diode laser brightness and the development of techniques for efficiently coupling multiple emitters. In this paper, we demonstrate further brightness improvement and power-scaling enabled by both the rise in chip brightness/power and the increase in number of chips coupled into a given numerical aperture. We report a new chip technology using x-REM design with brightness as high as 4.3 W/mm-mrad at a BPP of 3 mm-mrad. We also report record 315 W output from a 2×12 nLIGHT element with 105 μm diameter fiber using x-REM diodes and these diodes will allow next generation of fiber-coupled product capable of 250W output power from 105 μm/0.15 NA beam at 915 nm.

pH changes in frog rods upon manipulation of putative pH-regulating transport mechanisms.

PubMed

Kalamkarov, G; Pogozheva, I; Shevchenko, T; Koskelainen, A; Hemila, S; Donner, K

1996-10-01

Rod intracellular pH (pHi) in the intact frog retina was measured fluorometrically with the dye 2',7'-bis(2-carboxyethyl)-5(and-6)-carboxyfluorescein under treatments chosen to affect putative pH-regulating transport mechanisms in the plasma membrane. The purpose was to relate possible pHi changes to previously reported effects on photoresponses. In nominally bicarbonate-free Ringer, application of amiloride (1 mM) or substitution of 95 mM external Na+ by K+ or choline triggered monotonic but reversible acidifications, consistent with inhibition of Na+/H+ exchange. Bicarbonate-dependent mechanisms were characterized as follows: (1) Replacing half of a 12 mM phosphate buffer by bicarbonate caused a sustained rise of pHi. (2) Subsequent application of the anion transport inhibitor 4,4'-diisothiocyanatostilbene-2',2'-disulphonic acid (DIDS, 0.2 mM) set off a slow acidification. (3) Substitution of external Cl- by gluconate (95 mM) caused a rapid pHi rise both in normal Na+ and low-Na+ perfusion. (4) This effect was inhibited by DIDS. The results support a consistent explanation of parallel electrophysiological experiments on the assumption that intracellular acidifications reduce and alkalinizations (in a certain range) augment photoresponses. It is concluded that both Na+/H+ exchange and bicarbonate transport control rod pHi, modulating the light-sensitive current. Part of the bicarbonate transport is by Na(+)-independent HCO3-/Cl- exchange, but a further Na(+)-coupled bicarbonate import mechanism is implicated.

Coupling lattice Boltzmann and continuum equations for flow and reactive transport in porous media.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Coon, Ethan; Porter, Mark L.; Kang, Qinjun

2012-06-18

In spatially and temporally localized instances, capturing sub-reservoir scale information is necessary. Capturing sub-reservoir scale information everywhere is neither necessary, nor computationally possible. The lattice Boltzmann Method for solving pore-scale systems. At the pore-scale, LBM provides an extremely scalable, efficient way of solving Navier-Stokes equations on complex geometries. Coupling pore-scale and continuum scale systems via domain decomposition. By leveraging the interpolations implied by pore-scale and continuum scale discretizations, overlapping Schwartz domain decomposition is used to ensure continuity of pressure and flux. This approach is demonstrated on a fractured medium, in which Navier-Stokes equations are solved within the fracture while Darcy'smore » equation is solved away from the fracture Coupling reactive transport to pore-scale flow simulators allows hybrid approaches to be extended to solve multi-scale reactive transport.« less

Phosphorylation and transport in the Na-K-2Cl cotransporters, NKCC1 and NKCC2A, compared in HEK-293 cells.

PubMed

Hannemann, Anke; Flatman, Peter W

2011-03-25

Na-K-2Cl cotransporters help determine cell composition and volume. NKCC1 is widely distributed whilst NKCC2 is only found in the kidney where it plays a vital role reabsorbing 20% of filtered NaCl. NKCC2 regulation is poorly understood because of its restricted distribution and difficulties with its expression in mammalian cell cultures. Here we compare phosphorylation of the N-termini of the cotransporters, measured with phospho-specific antibodies, with bumetanide-sensitive transport of K(+) ((86)Rb(+)) (activity) in HEK-293 cells stably expressing fNKCC1 or fNKCC2A which were cloned from ferret kidney. Activities of transfected transporters were distinguished from those of endogenous ones by working at 37 °C. fNKCC1 and fNKCC2A activities were highest after pre-incubation of cells in hypotonic low-[Cl(-)] media to reduce cell [Cl(-)] and volume during flux measurement. Phosphorylation of both transporters more than doubled. Pre-incubation with ouabain also strongly stimulated fNKCC1 and fNKCC2A and substantially increased phosphorylation, whereas pre-incubation in Na(+)-free media maximally stimulated fNKCC1 and doubled its phosphorylation, but inhibited fNKCC2A, with a small increase in its phosphorylation. Kinase inhibitors halved phosphorylation and activity of both transporters whereas inhibition of phosphatases with calyculin A strongly increased phosphorylation of both transporters but only slightly stimulated fNKCC1 and inhibited fNCCC2A. Thus kinase inhibition reduced phosphorylation and transport, and transport stimulation was only seen when phosphorylation increased, but transport did not always increase with phosphorylation. This suggests phosphorylation of the N-termini determines the transporters' potential capacity to move ions, but final activity also depends on other factors. Transport cannot be reliably inferred solely using phospho-specific antibodies on whole-cell lysates.

Effect of diet containing phytate and phytase on the activity and messenger ribonucleic acid expression of carbohydrase and transporter in chickens.

PubMed

Liu, N; Ru, Y J; Li, F D; Cowieson, A J

2008-12-01

The effect of dietary phytate and phytase on carbohydrase activity and hexose transport was investigated in broiler chickens. Diets containing phytate P (2.2 or 4.4 g/kg) with different phytase dose rates (0, 500, or 1,000 phytase units/kg) were fed to 504 female Cobb chicks for 3 wk. Diets containing high phytate concentrations depressed (P < 0.05) BW and G:F, whereas phytase supplementation improved (P < 0.05) the performance of birds. In the duodenum, phytate decreased (P < 0.05) the activities of disaccharidases, Na(+)K(+)-ATPase, and glucose concentrations by 5 to 11%, but phytase enhanced (P < 0.05) the concentrations of amylase, sucrase, maltase, Na(+)K(+)-ATPase, and glucose by 5 to 30%. In the jejunum, phytate decreased (P < 0.05) the concentrations of amylase, sucrase, Na(+)K(+)-ATPase, and glucose by 10 to 22%, and phytase alleviated the negative effect of phytate on the above variables. Ingestion of diets containing phytate also decreased (P < 0.05) serum amylase activity and glucose concentration, and phytase enhanced (P < 0.05) serum concentrations of amylase, sucrase, maltase, Na(+)K(+)-ATPase, and glucose. There were also interactions (P < 0.05) between phytate and phytase on the concentrations of serum amylase, duodenal amylase, sucrase, and jejunal glucose. Enzymatic analysis at a molecular level showed that neither phytate nor phytase influenced the mRNA expression of sucrase-isomaltase in the small intestine. Also, the investigation into the sodium glucose cotransporter gene may challenge the mechanism by which phytate interferes with glucose utilization, as partly indicated by bird performance, and transmembrane transport because diets containing increased phytate upregulated (P < 0.05) the mRNA expression of the sodium glucose cotransporter gene in duodenum and did not influence it in the jejunum. These results indicate that phytate can impair endogenous carbohydrase activity and digestive competence, and phytase can ameliorate these effects

Development of a three-dimensional, regional, coupled wave, current, and sediment-transport model

USGS Publications Warehouse

Warner, J.C.; Sherwood, C.R.; Signell, R.P.; Harris, C.K.; Arango, H.G.

2008-01-01

We are developing a three-dimensional numerical model that implements algorithms for sediment transport and evolution of bottom morphology in the coastal-circulation model Regional Ocean Modeling System (ROMS v3.0), and provides a two-way link between ROMS and the wave model Simulating Waves in the Nearshore (SWAN) via the Model-Coupling Toolkit. The coupled model is applicable for fluvial, estuarine, shelf, and nearshore (surfzone) environments. Three-dimensional radiation-stress terms have been included in the momentum equations, along with effects of a surface wave roller model. The sediment-transport algorithms are implemented for an unlimited number of user-defined non-cohesive sediment classes. Each class has attributes of grain diameter, density, settling velocity, critical stress threshold for erosion, and erodibility constant. Suspended-sediment transport in the water column is computed with the same advection-diffusion algorithm used for all passive tracers and an additional algorithm for vertical settling that is not limited by the CFL criterion. Erosion and deposition are based on flux formulations. A multi-level bed framework tracks the distribution of every size class in each layer and stores bulk properties including layer thickness, porosity, and mass, allowing computation of bed morphology and stratigraphy. Also tracked are bed-surface properties including active-layer thickness, ripple geometry, and bed roughness. Bedload transport is calculated for mobile sediment classes in the top layer. Bottom-boundary layer submodels parameterize wave-current interactions that enhance bottom stresses and thereby facilitate sediment transport and increase bottom drag, creating a feedback to the circulation. The model is demonstrated in a series of simple test cases and a realistic application in Massachusetts Bay. 

Transport direction determines the kinetics of substrate transport by the glutamate transporter EAAC1

PubMed Central

Zhang, Zhou; Tao, Zhen; Gameiro, Armanda; Barcelona, Stephanie; Braams, Simona; Rauen, Thomas; Grewer, Christof

2007-01-01

Glutamate transport by the excitatory amino acid carrier EAAC1 is known to be reversible. Thus, glutamate can either be taken up into cells, or it can be released from cells through reverse transport, depending on the electrochemical gradient of the co- and countertransported ions. However, it is unknown how fast and by which reverse transport mechanism glutamate can be released from cells. Here, we determined the steady- and pre-steady-state kinetics of reverse glutamate transport with submillisecond time resolution. First, our results suggest that glutamate and Na+ dissociate from their cytoplasmic binding sites sequentially, with glutamate dissociating first, followed by the three cotransported Na+ ions. Second, the kinetics of glutamate transport depend strongly on transport direction, with reverse transport being faster but less voltage-dependent than forward transport. Third, electrogenicity is distributed over several reverse transport steps, including intracellular Na+ binding, reverse translocation, and reverse relocation of the K+-bound EAAC1. We propose a kinetic model, which is based on a “first-in-first-out” mechanism, suggesting that glutamate association, with its extracellular binding site as well as dissociation from its intracellular binding site, precedes association and dissociation of at least one Na+ ion. Our model can be used to predict rates of glutamate release from neurons under physiological and pathophysiological conditions. PMID:17991780

Contribution of Glucose Transport to the Control of the Glycolytic Flux in Trypanosoma brucei

NASA Astrophysics Data System (ADS)

Bakker, Barbara M.; Walsh, Michael C.; Ter Kuile, Benno H.; Mensonides, Femke I. C.; Michels, Paul A. M.; Opperdoes, Fred R.; Westerhoff, Hans V.

1999-08-01

The rate of glucose transport across the plasma membrane of the bloodstream form of Trypanosoma brucei was modulated by titration of the hexose transporter with the inhibitor phloretin, and the effect on the glycolytic flux was measured. A rapid glucose uptake assay was developed to measure the transport activity independently of the glycolytic flux. Phloretin proved a competitive inhibitor. When the effect of the intracellular glucose concentration on the inhibition was taken into account, the flux control coefficient of the glucose transporter was between 0.3 and 0.5 at 5 mM glucose. Because the flux control coefficients of all steps in a metabolic pathway sum to 1, this result proves that glucose transport is not the rate-limiting step of trypanosome glycolysis. Under physiological conditions, transport shares the control with other steps. At glucose concentrations much lower than physiological, the glucose carrier assumed all control, in close agreement with model predictions.

Dynamical coupling between magnetic equilibrium and transport in tokamak scenario modelling, with application to current ramps

NASA Astrophysics Data System (ADS)

Fable, E.; Angioni, C.; Ivanov, A. A.; Lackner, K.; Maj, O.; Medvedev, S. Yu; Pautasso, G.; Pereverzev, G. V.; Treutterer, W.; the ASDEX Upgrade Team

2013-07-01

The modelling of tokamak scenarios requires the simultaneous solution of both the time evolution of the plasma kinetic profiles and of the magnetic equilibrium. Their dynamical coupling involves additional complications, which are not present when the two physical problems are solved separately. Difficulties arise in maintaining consistency in the time evolution among quantities which appear in both the transport and the Grad-Shafranov equations, specifically the poloidal and toroidal magnetic fluxes as a function of each other and of the geometry. The required consistency can be obtained by means of iteration cycles, which are performed outside the equilibrium code and which can have different convergence properties depending on the chosen numerical scheme. When these external iterations are performed, the stability of the coupled system becomes a concern. In contrast, if these iterations are not performed, the coupled system is numerically stable, but can become physically inconsistent. By employing a novel scheme (Fable E et al 2012 Nucl. Fusion submitted), which ensures stability and physical consistency among the same quantities that appear in both the transport and magnetic equilibrium equations, a newly developed version of the ASTRA transport code (Pereverzev G V et al 1991 IPP Report 5/42), which is coupled to the SPIDER equilibrium code (Ivanov A A et al 2005 32nd EPS Conf. on Plasma Physics (Tarragona, 27 June-1 July) vol 29C (ECA) P-5.063), in both prescribed- and free-boundary modes is presented here for the first time. The ASTRA-SPIDER coupled system is then applied to the specific study of the modelling of controlled current ramp-up in ASDEX Upgrade discharges.

Evidence for coupled transport of bicarbonate and sodium in cultured bovine corneal endothelial cells.

PubMed

Jentsch, T J; Keller, S K; Koch, M; Wiederholt, M

1984-01-01

Using intracellular microelectrode technique, the response of the voltage V across the plasma membrane of cultured bovine corneal endothelial cells to changes in sodium and bicarbonate concentrations was investigated. (1) The electrical response to changes in [HCO3-]o (depolarization upon lowering and hyperpolarization upon raising [HCO3-]o) was dependent on sodium. Lithium could fairly well be substituted for sodium, whereas potassium or choline were much less effective. (2) Removal of external sodium caused a depolarization, while a readdition led to a hyperpolarization, which increased with time of preincubation in the sodium-depleted medium. (3) The response to changes in [Na+]o was dependent on bicarbonate. In a nominally bicarbonate-free medium, its amplitude was decreased or even reversed in sign. (4) Application of SITS or DIDS (10(-3) M) had a similar effect on the response to sodium as bicarbonate-depleted medium. (5) At [Na+]o = 151 mM and [HCO3-]o = 46 mM, the transients of V depended, with 39.0 +/- 9.0 (SD) mV/decade, on bicarbonate and, with 15.3 +/- 5.8 (SD) mV/decade, on sodium. (6) After the preincubation of cells with lithium, replacement of Li by choline led to similar effects as the replacement of sodium by choline, though the response of V was smaller with Li. This response could be reduced or reversed by the removal of bicarbonate or by the application of SITS. (7) Amiloride (10(-3) M) caused a reversible hyperpolarization of the steady-state potential by 8.5 +/- 2.6 mV (SD). It did not affect the immediate response to changes in [Na+]o or [HCO3-]o, but reduced the speed of regaining the steady-state potential after a change in [HCO3-]o. (8) Ouabain (10(-4) M) caused a fast depolarization of -6.8 +/- 1.1 (SD) mV, which was followed by a continuing slower depolarization. The effect was almost identical at 10(-5) M. (9) It is suggested, that corneal endothelial cells possess a cotransport for sodium and bicarbonate, which transports net negative

Na/sup +/-independent, phloretin-sensitive monosaccharide transport system in isolated intestinal epithelial cells. [Chickens

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kimmich, G.A.; Randles, J.

1975-01-01

A monosaccharide transport system in addition to the active Na/sup +/-dependent system characteristic of the brush border surface of vertebrate intestinal tissue has been identified in isolated chick intestinal epithelial cells. The newly described system differs in several characteristics from the Na/sup +/-dependent process, including function in the absence of Na/sup +/; a high sensitivity to phloretin, relative insensitivity to phlorizin; different substrate specificity; and a very high K/sub T/ and V/sub max/. The system apparently functions only in a facilitated diffusion manner so that it serves to move monosaccharide across the cell membrane down its chemical gradient. An appreciablemore » fraction of total sugar efflux occurs via the Na/sup +/-independent carrier from cells which have accumulated sugar to a steady state. Phloretin selectively blocks this efflux so that a normal steady-state sugar gradient of seven- to eightfold is transformed to a new steady-state gradient which is greater than 14-fold. Locus of the new system is tentatively ascribed to the serosal cell surface where it would serve for monosaccharide transfer between enterocyte and lamina propria of the villus. (auth)« less

Close Association of Carbonic Anhydrase (CA2a and CA15a), Na+/H+ Exchanger (Nhe3b), and Ammonia Transporter Rhcg1 in Zebrafish Ionocytes Responsible for Na+ Uptake

PubMed Central

Ito, Yusuke; Kobayashi, Sayako; Nakamura, Nobuhiro; Miyagi, Hisako; Esaki, Masahiro; Hoshijima, Kazuyuki; Hirose, Shigehisa

2013-01-01

Freshwater (FW) fishes actively absorb salt from their environment to tolerate low salinities. We previously reported that vacuolar-type H+-ATPase/mitochondrion-rich cells (H-MRCs) on the skin epithelium of zebrafish larvae (Danio rerio) are primary sites for Na+ uptake. In this study, in an attempt to clarify the mechanism for the Na+ uptake, we performed a systematic analysis of gene expression patterns of zebrafish carbonic anhydrase (CA) isoforms and found that, of 12 CA isoforms, CA2a and CA15a are highly expressed in H-MRCs at larval stages. The ca2a and ca15a mRNA expression were salinity-dependent; they were upregulated in 0.03 mM Na+ water whereas ca15a but not ca2a was down-regulated in 70 mM Na+ water. Immunohistochemistry demonstrated cytoplasmic distribution of CA2a and apical membrane localization of CA15a. Furthermore, cell surface immunofluorescence staining revealed external surface localization of CA15a. Depletion of either CA2a or CA15a expression by Morpholino antisense oligonucleotides resulted in a significant decrease in Na+ accumulation in H-MRCs. An in situ proximity ligation assay demonstrated a very close association of CA2a, CA15a, Na+/H+ exchanger 3b (Nhe3b), and Rhcg1 ammonia transporter in H-MRC. Our findings suggest that CA2a, CA15a, and Rhcg1 play a key role in Na+uptake under FW conditions by forming a transport metabolon with Nhe3b. PMID:23565095

Implications of Lagrangian transport for coupled chemistry-climate simulations

NASA Astrophysics Data System (ADS)

Stenke, A.; Dameris, M.; Grewe, V.; Garny, H.

2008-10-01

For the first time a purely Lagrangian transport algorithm is applied in a fully coupled chemistry-climate model (CCM). We use the Lagrangian scheme ATTILA for the transport of water vapour, cloud water and chemical trace species in the ECHAM4.L39(DLR)/CHEM (E39C) CCM. The advantage of the Lagrangian approach is that it is numerically non-diffusive and therefore maintains steeper and more realistic gradients than the operational semi-Lagrangian transport scheme. In case of radiatively active species changes in the simulated distributions feed back to model dynamics which in turn affect the modelled transport. The implications of the Lagrangian transport scheme for stratospheric model dynamics and tracer distributions in the upgraded model version E39C-ATTILA (E39C-A) are evaluated by comparison with observations and results of the E39C model with the operational semi-Lagrangian advection scheme. We find that several deficiencies in stratospheric dynamics in E39C seem to originate from a pronounced modelled wet bias and an associated cold bias in the extra-tropical lowermost stratosphere. The reduction of the simulated moisture and temperature bias in E39C-A leads to a significant advancement of stratospheric dynamics in terms of the mean state as well as annual and interannual variability. As a consequence of the favourable numerical characteristics of the Lagrangian transport scheme and the improved model dynamics, E39C-A generally shows more realistic stratospheric tracer distributions: Compared to E39C high stratospheric chlorine (Cly) concentrations extend further downward and agree now well with analyses derived from observations. Therefore E39C-A realistically covers the altitude of maximum ozone depletion in the stratosphere. The location of the ozonopause, i.e. the transition from low tropospheric to high stratospheric ozone values, is also clearly improved in E39C-A. Furthermore, the simulated temporal evolution of stratospheric Cly in the past is

Transport and mixing in strongly coupled dusty plasma medium

NASA Astrophysics Data System (ADS)

Dharodi, Vikram; Das, Amita; Patel, Bhavesh

2016-10-01

The generalized hydrodynamic (GHD) fluid model has been employed to study the transport and mixing properties of Dusty plasma medium in strong coupling limit. The response of lighter electron and ion species to the dust motion is taken to be instantaneous i.e. inertia-less. Thus the electron and ion density are presumed to follow the Boltzman relation. In the incompressible limit (i-GHD) the model supports Transverse Shear wave in contrast to the Hydrodynamic fluids. It has been shown that the presence of these waves leads to a better mixing of fluid in this case. Several cases of flow configuration have been considered for the study. The transport and mixing attributes have been quantified by studying the dynamical evolution of tracer particles in the system. The diffusion and clustering of these test particles are directly linked to the mixing characteristic of a medium. The displacement of these particles provides for a quantitative estimate of the diffusion coefficient of the medium. It is shown that these test particles often organize themselves in spatially inhomogeneous pattern leading to the phenomena of clustering.

Integrated Control of Na Transport along the Nephron

PubMed Central

Schnermann, Jürgen

2015-01-01

The kidney filters vast quantities of Na at the glomerulus but excretes a very small fraction of this Na in the final urine. Although almost every nephron segment participates in the reabsorption of Na in the normal kidney, the proximal segments (from the glomerulus to the macula densa) and the distal segments (past the macula densa) play different roles. The proximal tubule and the thick ascending limb of the loop of Henle interact with the filtration apparatus to deliver Na to the distal nephron at a rather constant rate. This involves regulation of both filtration and reabsorption through the processes of glomerulotubular balance and tubuloglomerular feedback. The more distal segments, including the distal convoluted tubule (DCT), connecting tubule, and collecting duct, regulate Na reabsorption to match the excretion with dietary intake. The relative amounts of Na reabsorbed in the DCT, which mainly reabsorbs NaCl, and by more downstream segments that exchange Na for K are variable, allowing the simultaneous regulation of both Na and K excretion. PMID:25098598

Structural basis of the alternating-access mechanism in a bile acid transporter

NASA Astrophysics Data System (ADS)

Zhou, Xiaoming; Levin, Elena J.; Pan, Yaping; McCoy, Jason G.; Sharma, Ruchika; Kloss, Brian; Bruni, Renato; Quick, Matthias; Zhou, Ming

2014-01-01

Bile acids are synthesized from cholesterol in hepatocytes and secreted through the biliary tract into the small intestine, where they aid in absorption of lipids and fat-soluble vitamins. Through a process known as enterohepatic recirculation, more than 90% of secreted bile acids are then retrieved from the intestine and returned to the liver for resecretion. In humans, there are two Na+-dependent bile acid transporters involved in enterohepatic recirculation, the Na+-taurocholate co-transporting polypeptide (NTCP; also known as SLC10A1) expressed in hepatocytes, and the apical sodium-dependent bile acid transporter (ASBT; also known as SLC10A2) expressed on enterocytes in the terminal ileum. In recent years, ASBT has attracted much interest as a potential drug target for treatment of hypercholesterolaemia, because inhibition of ASBT reduces reabsorption of bile acids, thus increasing bile acid synthesis and consequently cholesterol consumption. However, a lack of three-dimensional structures of bile acid transporters hampers our ability to understand the molecular mechanisms of substrate selectivity and transport, and to interpret the wealth of existing functional data. The crystal structure of an ASBT homologue from Neisseria meningitidis (ASBTNM) in detergent was reported recently, showing the protein in an inward-open conformation bound to two Na+ and a taurocholic acid. However, the structural changes that bring bile acid and Na+ across the membrane are difficult to infer from a single structure. To understand the structural changes associated with the coupled transport of Na+ and bile acids, here we solved two structures of an ASBT homologue from Yersinia frederiksenii (ASBTYf) in a lipid environment, which reveal that a large rigid-body rotation of a substrate-binding domain gives the conserved `crossover' region, where two discontinuous helices cross each other, alternating accessibility from either side of the cell membrane. This result has implications

Structural basis of the alternating-access mechanism in a bile acid transporter

PubMed Central

Zhou, Xiaoming; Levin, Elena J.; Pan, Yaping; McCoy, Jason G.; Sharma, Ruchika; Kloss, Brian; Bruni, Renato; Quick, Matthias; Zhou, Ming

2014-01-01

Bile acids are synthesized from cholesterol in hepatocytes and secreted via the biliary tract into the small intestine, where they aid in absorption of lipids and fat-soluble vitamins. Through a process known as enterohepatic recirculation, more than 90% of secreted bile acids are then retrieved from the intestine and returned to the liver for re-secretion1. In humans, there are two Na+-dependent bile acid transporters involved in enterohepatic recirculation, the Na+-taurocholate co-transporting polypeptide (NTCP or SLC10A1) expressed in hepatocytes, and the apical sodium-dependent bile acid transporter (ASBT or SLC10A2) expressed on enterocytes in the terminal ileum2. In recent years, ASBT has attracted much interest as a potential drug target for treatment of hypercholesterolemia, because inhibition of ASBT reduces reabsorption of bile acids, thus increasing bile acid synthesis and consequently cholesterol consumption3,4. However, a lack of 3-dimensional structures of bile acid transporters hampers our ability to understand the molecular mechanisms of substrate selectivity and transport, and to interpret the wealth of existing functional data2,5-8. The crystal structure of an ASBT homolog from Neisseria meningitidis (ASBTNM) in detergent was reported recently9, showing the protein in an inward-open conformation bound to two Na+ and a taurocholic acid. However, the structural changes that bring bile acid and Na+ across the membrane are difficult to infer from a single structure. To understand better the structural changes associated with the coupled transport of Na+ and bile acids, we crystallized and solved two structures of a ASBT homolog from Yersinia frederiksenii (ASBTYf) in a lipid environment, which reveal that a large rigid-body rotation of a substrate-binding domain gives alternate accessibility to the highly conserved “crossover” region, where two discontinuous transmembrane helices cross each other. This result has implications for the location and

Characterization of an AtCCX5 gene from Arabidopsis thaliana that involves in high-affinity K{sup +} uptake and Na{sup +} transport in yeast

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Xinxin; Zhang, Min; Takano, Tetsuo

Highlights: {yields} The AtCCX5 protein coding a putative cation calcium exchanger was characterized. {yields} AtCCX5 expressed in yeast was localized in the plasma membrane and nuclear periphery. {yields} AtCCX5 protein did not show the same transport properties as the CAXs. {yields} AtCCX5 protein involves in mediating high-affinity K{sup +} uptake in yeast. {yields} AtCCX5 protein also involves in Na{sup +} transport in yeast. -- Abstract: The gene for a putative cation calcium exchanger (CCX) from Arabidopsis thaliana, AtCCX5, was cloned and its function was analyzed in yeast. Green fluorescent protein-tagged AtCCX5 expressed in yeast was localized in the plasma membranemore » and nuclear periphery. The yeast transformants expressing AtCCX5 were created and their growth in the presence of various cations (K{sup +}, Na{sup +}, Ca{sup 2+}, Mg{sup 2+}, Fe{sup 2+}, Cu{sup 2+}, Co{sup 2+}, Cd{sup 2+}, Mn{sup 2+}, Ba{sup 2+}, Ni{sup 2+}, Zn{sup 2+}, and Li{sup +}) were analyzed. AtCCX5 expression was found to affect the response to K{sup +} and Na{sup +} in yeast. The AtCCX5 transformant also showed a little better growth to Zn{sup 2+}. The yeast mutant 9.3 expressing AtCCX5 restored growth of the mutant on medium with low K{sup +} (0.5 mM), and also suppressed its Na{sup +} sensitivity. Ion uptake experiments showed that AtCCX5 mediated relatively high-affinity K{sup +} uptake and was also involved in Na{sup +} transport in yeast. Taken together, these findings suggest that the AtCCX5 is a novel transport protein involves in mediating high-affinity K{sup +} uptake and Na{sup +} transport in yeast.« less

Mechanism of the Association between Na+ Binding and Conformations at the Intracellular Gate in Neurotransmitter:Sodium Symporters*

PubMed Central

Stolzenberg, Sebastian; Quick, Matthias; Zhao, Chunfeng; Gotfryd, Kamil; Khelashvili, George; Gether, Ulrik; Loland, Claus J.; Javitch, Jonathan A.; Noskov, Sergei; Weinstein, Harel; Shi, Lei

2015-01-01

Neurotransmitter:sodium symporters (NSSs) terminate neurotransmission by Na+-dependent reuptake of released neurotransmitters. Previous studies suggested that Na+-binding reconfigures dynamically coupled structural elements in an allosteric interaction network (AIN) responsible for function-related conformational changes, but the intramolecular pathway of this mechanism has remained uncharted. We describe a new approach for the modeling and analysis of intramolecular dynamics in the bacterial NSS homolog LeuT. From microsecond-scale molecular dynamics simulations and cognate experimental verifications in both LeuT and human dopamine transporter (hDAT), we apply the novel method to identify the composition and the dynamic properties of their conserved AIN. In LeuT, two different perturbations disrupting Na+ binding and transport (i.e. replacing Na+ with Li+ or the Y268A mutation at the intracellular gate) affect the AIN in strikingly similar ways. In contrast, other mutations that affect the intracellular gate (i.e. R5A and D369A) do not significantly impair Na+ cooperativity and transport. Our analysis shows these perturbations to have much lesser effects on the AIN, underscoring the sensitivity of this novel method to the mechanistic nature of the perturbation. Notably, this set of observations holds as well for hDAT, where the aligned Y335A, R60A, and D436A mutations also produce different impacts on Na+ dependence. Thus, the detailed AIN generated from our method is shown to connect Na+ binding with global conformational changes that are critical for the transport mechanism. That the AIN between the Na+ binding sites and the intracellular gate in bacterial LeuT resembles that in eukaryotic hDAT highlights the conservation of allosteric pathways underlying NSS function. PMID:25869126

Mechanism of the association between Na + binding and conformations at the intracellular gate in neurotransmitter:sodium symporters

DOE PAGES

Stolzenberg, Sebastian; Quick, Matthias; Zhao, Chunfeng; ...

2015-04-13

Neurotransmitter:sodium symporters (NSSs) terminate neurotransmission by Na +-dependent reuptake of released neurotransmitters. Previous studies suggested that Na +-binding reconfigures dynamically coupled structural elements in an allosteric interaction network (AIN) responsible for function-related conformational changes, but the intramolecular pathway of this mechanism has remained uncharted. Here we describe a new approach for the modeling and analysis of intramolecular dynamics in the bacterial NSS homolog LeuT. From microsecond-scale molecular dynamics simulations and cognate experimental verifications in both LeuT and human dopamine transporter (hDAT), we apply the novel method to identify the composition and the dynamic properties of their conserved AIN. In LeuT,more » two different perturbations disrupting Na+ binding and transport ( i.e. replacing Na + with Li + or the Y268A mutation at the intracellular gate) affect the AIN in strikingly similar ways. In contrast, other mutations that affect the intracellular gate (i.e. R5A and D369A) do not significantly impair Na + cooperativity and transport. Our analysis shows these perturbations to have much lesser effects on the AIN, underscoring the sensitivity of this novel method to the mechanistic nature of the perturbation. Notably, this set of observations holds as well for hDAT, where the aligned Y335A, R60A, and D436A mutations also produce different impacts on Na + dependence. Furthermore, the detailed AIN generated from our method is shown to connect Na + binding with global conformational changes that are critical for the transport mechanism. Lastly, that the AIN between the Na + binding sites and the intracellular gate in bacterial LeuT resembles that in eukaryotic hDAT highlights the conservation of allosteric pathways underlying NSS function.« less

A coupled hydrodynamic-hydrochemical modeling for predicting mineral transport in a natural acid drainage system.

NASA Astrophysics Data System (ADS)

Zegers Risopatron, G., Sr.; Navarro, L.; Montserrat, S., Sr.; McPhee, J. P.; Niño, Y.

2017-12-01

The geochemistry of water and sediments, coupled with hydrodynamic transport in mountainous channels, is of particular interest in central Chilean Andes due to natural occurrence of acid waters. In this paper, we present a coupled transport and geochemical model to estimate and understand transport processes and fate of minerals at the Yerba Loca Basin, located near Santiago, Chile. In the upper zone, water presentes low pH ( 3) and high concentrations of iron, aluminum, copper, manganese and zinc. Acidity and minerals are the consequence of water-rock interactions in hydrothermal alteration zones, rich in sulphides and sulphates, covered by seasonal snow and glaciers. Downstream, as a consequence of neutral to alkaline lateral water contributions (pH >7) along the river, pH increases and concentration of solutes decreases. The mineral transport model has three components: (i) a hydrodynamic model, where we use HEC-RAS to solve 1D Saint-Venant equations, (ii) a sediment transport model to estimate erosion and sedimentation rates, which quantify minerals transference between water and riverbed and (iii) a solute transport model, based on the 1D OTIS model which takes into account the temporal delay in solutes transport that typically is observed in natural channels (transient storage). Hydrochemistry is solved using PHREEQC, a software for speciation and batch reaction. Our results show that correlation between mineral precipitation and dissolution according to pH values changes along the river. Based on pH measurements (and according to literature) we inferred that main minerals in the water system are brochantite, ferrihydrite, hydrobasaluminite and schwertmannite. Results show that our model can predict the transport and fate of minerals and metals in the Yerba Loca Basin. Mineral dissolution and precipitation process occur for limited ranges of pH values. When pH values are increased, iron minerals (schwertmannite) are the first to precipitate ( 2.5

Derivation of Hodgkin-Huxley equations for a Na+ channel from a master equation for coupled activation and inactivation

NASA Astrophysics Data System (ADS)

Vaccaro, S. R.

2016-11-01

The Na+ current in nerve and muscle membranes may be described in terms of the activation variable m (t ) and the inactivation variable h (t ) , which are dependent on the transitions of S4 sensors of each of the Na+ channel domains DI to DIV. The time-dependence of the Na+ current and the rate equations satisfied by m (t ) and h (t ) may be derived from the solution to a master equation that describes the coupling between two or three activation sensors regulating the Na+ channel conductance and a two-stage inactivation process. If the inactivation rate from the closed or open states increases as the S4 sensors activate, a more general form of the Hodgkin-Huxley expression for the open-state probability may be derived where m (t ) is dependent on both activation and inactivation processes. The voltage dependence of the rate functions for inactivation and recovery from inactivation are consistent with the empirically determined expressions and exhibit saturation for both depolarized and hyperpolarized clamp potentials.

Coupling between Transport and Injection Properties of Pentacene Field-Effect Transistors with Different Morphologies

NASA Astrophysics Data System (ADS)

Lee, Keanchuan; Weis, Martin; Taguchi, Dai; Manaka, Takaaki; Iwamoto, Mitsumasa

2013-08-01

We investigated the injection and transport properties of pentacene organic field-effect transistors (OFETs) with inclined and lamellar pentacene grains at various mutual ratios. Although the threshold voltage was conserved and no additional trapping on grain boundaries was suggested from the current-voltage measurements, the contact resistance and mobility increased linearly with the lamellar phase content. We showed that a model based on the coupling between both transport and injection properties via a space charge field caused by injected and trapped carriers accounts for these results.

Development of a 1.5D plasma transport code for coupling to full orbit runaway electron simulations

NASA Astrophysics Data System (ADS)

Lore, J. D.; Del Castillo-Negrete, D.; Baylor, L.; Carbajal, L.

2017-10-01

A 1.5D (1D radial transport + 2D equilibrium geometry) plasma transport code is being developed to simulate runaway electron generation, mitigation, and avoidance by coupling to the full-orbit kinetic electron transport code KORC. The 1.5D code solves the time-dependent 1D flux surface averaged transport equations with sources for plasma density, pressure, and poloidal magnetic flux, along with the Grad-Shafranov equilibrium equation for the 2D flux surface geometry. Disruption mitigation is simulated by introducing an impurity neutral gas `pellet', with impurity densities and electron cooling calculated from ionization, recombination, and line emission rate coefficients. Rapid cooling of the electrons increases the resistivity, inducing an electric field which can be used as an input to KORC. The runaway electron current is then included in the parallel Ohm's law in the transport equations. The 1.5D solver will act as a driver for coupled simulations to model effects such as timescales for thermal quench, runaway electron generation, and pellet impurity mixtures for runaway avoidance. Current progress on the code and details of the numerical algorithms will be presented. Work supported by the US DOE under DE-AC05-00OR22725.

Immunolocalization of chloride transporters to gill epithelia of euryhaline teleosts with opposite salinity-induced Na+/K+-ATPase responses.

PubMed

Tang, Cheng-Hao; Hwang, Lie-Yueh; Shen, I-Da; Chiu, Yu-Hui; Lee, Tsung-Han

2011-12-01

Opposite patterns of branchial Na(+)/K(+)-ATPase (NKA) responses were found in euryhaline milkfish (Chanos chanos) and pufferfish (Tetraodon nigroviridis) upon salinity challenge. Because the electrochemical gradient established by NKA is thought to be the driving force for transcellular Cl(-) transport in fish gills, the aim of this study was to explore whether the differential patterns of NKA responses found in milkfish and pufferfish would lead to distinct distribution of Cl(-) transporters in their gill epithelial cells indicating different Cl(-) transport mechanisms. In this study, immunolocalization of various Cl(-) transport proteins, including Na(+)/K(+)/2Cl(-) cotransporter (NKCC), cystic fibrosis transmembrane conductance regulator (CFTR), anion exchanger 1 (AE1), and chloride channel 3 (ClC-3), were double stained with NKA, the basolateral marker of branchial mitochondrion-rich cells (MRCs), to reveal the localization of these transporter proteins in gill MRC of FW- or SW-acclimated milkfish and pufferfish. Confocal microscopic observations showed that the localization of these transport proteins in the gill MRCs of the two studied species were similar. However, the number of gill NKA-immunoreactive (IR) cells in milkfish and pufferfish exhibited to vary with environmental salinities. An increase in the number of NKA-IR cells should lead to the elevation of NKA activity in FW milkfish and SW pufferfish. Taken together, the opposite branchial NKA responses observed in milkfish and pufferfish upon salinity challenge could be attributed to alterations in the number of NKA-IR cells. Furthermore, the localization of these Cl(-) transporters in gill MRCs of the two studied species was identical. It depicted the two studied euryhaline species possess the similar Cl(-) transport mechanisms in gills.

Neurotransmitter and psychostimulant recognition by the dopamine transporter

PubMed Central

Wang, Kevin H.; Penmatsa, Aravind; Gouaux, Eric

2015-01-01

Na+/Cl−-coupled biogenic amine transporters are the primary targets of therapeutic and abused drugs, ranging from antidepressants to the psychostimulants cocaine and amphetamines, and to their cognate substrates. Here we determine x-ray crystal structures of the Drosophila melanogaster dopamine transporter (dDAT) bound to its substrate dopamine (DA), a substrate analogue 3,4-dichlorophenethylamine, the psychostimulants D-amphetamine, methamphetamine, or to cocaine and cocaine analogues. All ligands bind to the central binding site, located approximately halfway across the membrane bilayer, in close proximity to bound sodium and chloride ions. The central binding site recognizes three chemically distinct classes of ligands via conformational changes that accommodate varying sizes and shapes, thus illustrating molecular principles that distinguish substrates from inhibitors in biogenic amine transporters. PMID:25970245

Sodium-potassium-adenosinetriphosphatase-dependent sodium transport in the kidney: hormonal control.

PubMed

Féraille, E; Doucet, A

2001-01-01

Tubular reabsorption of filtered sodium is quantitatively the main contribution of kidneys to salt and water homeostasis. The transcellular reabsorption of sodium proceeds by a two-step mechanism: Na(+)-K(+)-ATPase-energized basolateral active extrusion of sodium permits passive apical entry through various sodium transport systems. In the past 15 years, most of the renal sodium transport systems (Na(+)-K(+)-ATPase, channels, cotransporters, and exchangers) have been characterized at a molecular level. Coupled to the methods developed during the 1965-1985 decades to circumvent kidney heterogeneity and analyze sodium transport at the level of single nephron segments, cloning of the transporters allowed us to move our understanding of hormone regulation of sodium transport from a cellular to a molecular level. The main purpose of this review is to analyze how molecular events at the transporter level account for the physiological changes in tubular handling of sodium promoted by hormones. In recent years, it also became obvious that intracellular signaling pathways interacted with each other, leading to synergisms or antagonisms. A second aim of this review is therefore to analyze the integrated network of signaling pathways underlying hormone action. Given the central role of Na(+)-K(+)-ATPase in sodium reabsorption, the first part of this review focuses on its structural and functional properties, with a special mention of the specificity of Na(+)-K(+)-ATPase expressed in renal tubule. In a second part, the general mechanisms of hormone signaling are briefly introduced before a more detailed discussion of the nephron segment-specific expression of hormone receptors and signaling pathways. The three following parts integrate the molecular and physiological aspects of the hormonal regulation of sodium transport processes in three nephron segments: the proximal tubule, the thick ascending limb of Henle's loop, and the collecting duct.

Inhibition of Na+ transport in lung epithelial cells by respiratory syncytial virus infection.

PubMed

Chen, Lan; Song, Weifeng; Davis, Ian C; Shrestha, Kedar; Schwiebert, Erik; Sullender, Wayne M; Matalon, Sadis

2009-05-01

We investigated the mechanisms by which respiratory syncytial virus (RSV) infection decreases vectorial Na+ transport across respiratory epithelial cells. Mouse tracheal epithelial (MTE) cells from either BALB/c or C57BL/6 mice and human airway H441 cells were grown on semipermeable supports under an air-liquid interface. Cells were infected with RSV-A2 and mounted in Ussing chambers for measurements of short-circuit currents (I(sc)). Infection with RSV for 24 hours (multiplicity of infection = 1) resulted in positive immunofluorescence for RSV antigen in less than 10% of MTE or H441 cells. In spite of the limited number of cells infected, RSV reduced both basal and amiloride-sensitive I(sc) in both MTE and H441 cells by approximately 50%, without causing a concomitant reduction in transepithelial resistance. Agents that increased intracellular cAMP (forskolin, cpt-CAMP, and IBMX) increased mainly Cl(-) secretion in MTE cells and Na+ absorption in H441 cells. RSV infection for 24 hours blunted both variables. In contrast, ouabain sensitive I(sc), measured across apically permeabilized H441 monolayers, remained unchanged. Western blot analysis of H441 cell lysates demonstrated reductions in alpha- but not gamma-ENaC subunit protein levels at 24 hours after RSV infection. The reduction in amiloride-sensitive I(sc) in H441 cells was prevented by pretreatment with inhibitors of de novo pyrimidine or purine synthesis (A77-1726 and 6-MP, respectively, 50 microM). Our results suggest that infection of both murine and human respiratory epithelial cells with RSV inhibits vectorial Na+ transport via nucleotide release. These findings are consistent with our previous studies showing reduced alveolar fluid clearance after RSV infection of BALB/c mice.

Characteristics of injury and recovery of net NO3- transport of barley seedlings from treatments of NaCl

NASA Technical Reports Server (NTRS)

Klobus, G.; Ward, M. R.; Huffaker, R. C.

1988-01-01

The nature of the injury and recovery of nitrate uptake (net uptake) from NaCl stress in young barley (Hordeum vulgare L, var CM 72) seedlings was investigated. Nitrate uptake was inhibited rapidly by NaCl, within 1 minute after exposure to 200 millimolar NaCl. The duration of exposure to saline conditions determined the time of recovery of NO3- uptake from NaCl stress. Recovery was dependent on the presence of NO3- and was inhibited by cycloheximide, 6-methylpurine, and cerulenin, respective inhibitors of protein, RNA, and sterol/fatty acid synthesis. These inhibitors also prevented the induction of the NO3- uptake system in uninduced seedlings. Uninduced seedlings exhibited endogenous NO3- transport activity that appeared to be constitutive. This constitutive activity was also inhibited by NaCl. Recovery of constitutive NO3- uptake did not require the presence of NO3-.

A novel Na+/HCO3--codependent choline transporter in the syncytial epithelium of the cestode Hymenolepis diminuta.

PubMed

Webb, R A; Xue, L

1998-02-01

Absorption of exogenous choline by the cestode Hymenolepis diminuta was found to be both Na+- and HCO3--dependent and, at pH 6 to 7, accounted for up to 65% of the total choline uptake. Na+/HCO3- dependent choline uptake was activated at approximately 6 mM HCO3- (EC50 approximately 9 mM), and, above 100 mM Na+, the rate of uptake was directly proportional to the Na+ concentration. Atempts to uncouple Na+-dependent uptake from HCO3--dependent uptake were not successful: K+-depolarization was without effect on HCO3--dependent choline uptake, and use of valinoomycin to hyperpolarize the brush-border membrane resulted in inhibition of uptake. Na-/HCO3--dependent choline uptake was not associated with solvent drag. The Na+/HCO3--dependent choline uptake displayed a Q10 of 6.4 (27 degrees to 37 degrees) and a relatively high activation energy of 126 kJ x mol(-1). At pH 6.0 and 7.0, Na-/HCO3--dependent choline uptake rates were similar, but Na+/HCO3--dependent choline uptake was reduced at pH 5.0. The Na+/HCO3--dependent choline uptake, at pH 7.0, displayed a Kt of approximately 500 microM and a Vmax of 4.01 pmol x mg wet weight(-1) x min(-1). The Na+/HCO3--dependent choline uptake was hemicholinium-3 sensitive, but not significantly inhibited by 200 microM bumetanide, 100 microM amiloride, benzamil, or EIPA or by 1 mM 4,4'-diisothiocyano-2,2'-stilbene disulfonate (DIDS) or 4-acetamido-4'-isothiocvanostilbene-2,2'-disulfonic acid (SITS). Although it remains to be shown that HCO3- uptake is coupled directly to both choline and Na+ uptake, the data suggest that choline up take occurs via choline/Na+/HCO3--co-trans porter.

Numerical modeling of coupled variably saturated fluid flow and reactive transport with fast and slow chemical reactions

NASA Astrophysics Data System (ADS)

Yeh, Gour-Tsyh (George); Siegel, Malcolm D.; Li, Ming-Hsu

2001-02-01

The couplings among chemical reaction rates, advective and diffusive transport in fractured media or soils, and changes in hydraulic properties due to precipitation and dissolution within fractures and in rock matrix are important for both nuclear waste disposal and remediation of contaminated sites. This paper describes the development and application of LEHGC2.0, a mechanistically based numerical model for simulation of coupled fluid flow and reactive chemical transport, including both fast and slow reactions in variably saturated media. Theoretical bases and numerical implementations are summarized, and two example problems are demonstrated. The first example deals with the effect of precipitation/dissolution on fluid flow and matrix diffusion in a two-dimensional fractured media. Because of the precipitation and decreased diffusion of solute from the fracture into the matrix, retardation in the fractured medium is not as large as the case wherein interactions between chemical reactions and transport are not considered. The second example focuses on a complicated but realistic advective-dispersive-reactive transport problem. This example exemplifies the need for innovative numerical algorithms to solve problems involving stiff geochemical reactions.

The serotonin transporter: Examination of the changes in transporter affinity induced by ligand binding

DOE Office of Scientific and Technical Information (OSTI.GOV)

Humphreys, C.J.

1989-01-01

The plasmalemmal serotonin transporter uses transmembrane gradients of Na{sup +}, Cl{sup {minus}} and K{sup +} to accumulate serotonin within blood platelets. Transport is competitively inhibited by the antidepressant imipramine. Like serotonin transport, imipramine binding requires Na{sup +}. Unlike serotonin, however, imipramine does not appear to be transported. To gain insight into the mechanism of serotonin transport the author have analyzed the influences of Na{sup +} and Cl{sup {minus}}, the two ions cotransported with serotonin, on both serotonin transport and the interaction of imipramine and other antidepressant drugs with the plasmalemmal serotonin transporter of human platelets. Additionally, the author have synthesized,more » purified and characterized the binding of 2-iodoimipramine to the serotonin transporter. Finally, the author have conducted a preliminary study of the inhibition of serotonin transport and imipramine binding produced by dicyclohexylcarbodiimide. My results reveal many instances of positive heterotropic cooperativity in ligand binding to the serotonin transporter. Na{sup +} binding enhances the transporters affinity for imipramine and several other antidepressant drugs, and also increases the affinity for Cl{sup {minus}}. Cl{sup {minus}} enhances the transporters affinity for imipramine, as well as for Na{sup +}. At concentrations in the range of its K{sub M} for transport serotonin is a competitive inhibitor of imipramine binding. At much higher concentrations, however, serotonin also inhibits imipramines dissociation rate constant. This latter effect which is Na{sup +}-independent and species specific, is apparently produced by serotonin binding at a second, low affinity site on, or near, the transporter complex. Iodoimipramine competitively inhibit both ({sup 3}H)imipramine binding and ({sup 3}H)serotonin transport.« less

Transport by SLC5A8 with subsequent inhibition of histone deacetylase 1 (HDAC1) and HDAC3 underlies the antitumor activity of 3-bromopyruvate.

PubMed

Thangaraju, Muthusamy; Karunakaran, Senthil K; Itagaki, Shiro; Gopal, Elangovan; Elangovan, Selvakumar; Prasad, Puttur D; Ganapathy, Vadivel

2009-10-15

3-bromopyruvate is an alkylating agent with antitumor activity. It is currently believed that blockade of adenosine triphosphate production from glycolysis and mitochondria is the primary mechanism responsible for this antitumor effect. The current studies uncovered a new and novel mechanism for the antitumor activity of 3-bromopyruvate. The transport of 3-bromopyruvate by sodium-coupled monocarboxylate transporter SMCT1 (SLC5A8), a tumor suppressor and a sodium (Na+)-coupled, electrogenic transporter for short-chain monocarboxylates, was studied using a mammalian cell expression and the Xenopus laevis oocyte expression systems. The effect of 3-bromopyruvate on histone deacetylases (HDACs) was monitored using the lysate of the human breast cancer cell line MCF7 and human recombinant HDAC isoforms as the enzyme sources. Cell viability was monitored by fluorescence-activated cell-sorting analysis and colony-formation assay. The acetylation status of histone H4 was evaluated by Western blot analysis. 3-Bromopyruvate is a transportable substrate for SLC5A8, and that transport process is Na+-coupled and electrogenic. MCF7 cells did not express SLC5A8 and were not affected by 3-bromopyruvate. However, when transfected with SLC5A8 or treated with inhibitors of DNA methylation, these cells underwent apoptosis in the presence of 3-bromopyruvate. This cell death was associated with the inhibition of HDAC1/HDAC3. Studies with different isoforms of human recombinant HDACs identified HDAC1 and HDAC3 as the targets for 3-bromopyruvate. 3-Bromopyruvate was transported into cells actively through the tumor suppressor SLC5A8, and the process was energized by an electrochemical Na+ gradient. Ectopic expression of the transporter in MCF7 cells led to apoptosis, and the mechanism involved the inhibition of HDAC1/HDAC3. Copyright (c) 2009 American Cancer Society.

Fully coupled approach to modeling shallow water flow, sediment transport, and bed evolution in rivers

NASA Astrophysics Data System (ADS)

Li, Shuangcai; Duffy, Christopher J.

2011-03-01

Our ability to predict complex environmental fluid flow and transport hinges on accurate and efficient simulations of multiple physical phenomenon operating simultaneously over a wide range of spatial and temporal scales, including overbank floods, coastal storm surge events, drying and wetting bed conditions, and simultaneous bed form evolution. This research implements a fully coupled strategy for solving shallow water hydrodynamics, sediment transport, and morphological bed evolution in rivers and floodplains (PIHM_Hydro) and applies the model to field and laboratory experiments that cover a wide range of spatial and temporal scales. The model uses a standard upwind finite volume method and Roe's approximate Riemann solver for unstructured grids. A multidimensional linear reconstruction and slope limiter are implemented, achieving second-order spatial accuracy. Model efficiency and stability are treated using an explicit-implicit method for temporal discretization with operator splitting. Laboratory-and field-scale experiments were compiled where coupled processes across a range of scales were observed and where higher-order spatial and temporal accuracy might be needed for accurate and efficient solutions. These experiments demonstrate the ability of the fully coupled strategy in capturing dynamics of field-scale flood waves and small-scale drying-wetting processes.

Ion pump as Brownian motor: theory of electroconformational coupling and proof of ratchet mechanism for Na,K-ATPase action

NASA Astrophysics Data System (ADS)

Tsong, Tian Yow; Chang, Cheng-Hung

2003-04-01

This article reviews some concepts of the Brownian Ratchet which are relevant to our discussion of mechanisms of action of Na,K-ATPase, a universal ion pump and an elemental motor protein of the biological cell. Under wide ranges of ionic compositions it can hydrolyze an ATP and use the γ-phosphorous bond energy of ATP to pump 3 Na + out of, and 2 K + into the cell, both being uphill transport. During the ATP-dependent pump cycle, the enzyme oscillates between E1 and E2 states. Our experiment replaces ATP with externally applied electric field of various waveforms, amplitudes, and frequencies. The field enforced-oscillation, or fluctuation of E1 and E2 states enables the enzyme to harvest energy from the applied field and convert it to the chemical gradient energy of cations. A theory of electroconformational coupling (TEC), which embodies all the essential features of the Brownian Ratchet, successfully simulates these experimental results. Our analysis based on a four-state TEC model indicates that the equilibrium and the rate constants of the transport system define the frequency and the amplitude of the field for the optimal activation. Waveform, frequency, and amplitude are three elements of signal. Thus, electric signal of the ion pump is found by TEC analysis of the experimental data. Electric noise (white) superimposed on an electric signal changes the pump efficiency and produces effects similar to the stochastic resonance reported in other biological systems. The TEC concept is compared with the most commonly used Michaelis-Menten enzyme mechanism (MME) for similarities and differences. Both MME and TEC are catalytic wheels, which recycle the catalyst in each turnover. However, a MME can only catalyze reaction of descending free energy while a TEC enzyme can catalyze reaction of ascending free energy by harvesting needed energy from an off-equilibrium electric noise. The TEC mechanism is shown to be applicable to other biological motors and engines, as

Enzymatic properties of separated isozymes of the Na,K-ATPase. Substrate affinities, kinetic cooperativity, and ion transport stoichiometry.

PubMed

Sweadner, K J

1985-09-25

There are two isozymes of the Na,K-ATPase, which can be purified separately from rat renal medulla and brainstem axolemma. Here the basic kinetic properties of the two Na,K-ATPases have been compared in conditions permitting enzyme turnover. The two isozymes are half-maximally activated at different concentrations of ATP, the axolemma Na,K-ATPase having the higher affinity. They are half-maximally activated by Na+ and K+ at very similar concentrations but show differences in cooperativity toward Na+. The affinities of both isozymes for ATP and Na+ are affected in a qualitatively similar way by variations in the concentration of K+. Both isozymes transport 22Na+ and 42K+ in a ratio close to 3:2 in artificial lipid vesicles. The two isozymes differ most strikingly in the inhibition of ATPase activity by ouabain. The axolemma Na,K-ATPase has a high affinity for ouabain with positive cooperativity, while the renal medulla Na,K-ATPase has a lower affinity with negative cooperativity. It is likely that the cooperativity differences are due to kinetic effects, reflecting different rates of conformation transitions during enzyme turnover. The functional result of the contrasting cooperativities is that the difference in sensitivity to ouabain is amplified.

Numerical modeling of coupled water flow and heat transport in soil and snow

Treesearch

Thijs J. Kelleners; Jeremy Koonce; Rose Shillito; Jelle Dijkema; Markus Berli; Michael H. Young; John M. Frank; William Massman

2016-01-01

A one-dimensional vertical numerical model for coupled water flow and heat transport in soil and snow was modified to include all three phases of water: vapor, liquid, and ice. The top boundary condition in the model is driven by incoming precipitation and the surface energy balance. The model was applied to three different terrestrial systems: A warm desert bare...

Temperature optimum of insulin-stimulated 2-deoxy-D-glucose uptake in rat adipocytes. Correlation of cellular transport with membrane spin-label and fluorescence-label data.

PubMed Central

Hyslop, P A; Kuhn, C E; Sauerheber, R D

1984-01-01

The effects of temperature alterations between 22 degrees C and 48 degrees C on basal and insulin-stimulated 2-deoxy-D-[1-14C]glucose uptake were examined in isolated rat adipocytes. A distinct optimum was found near physiological temperature for uptake in the presence of maximally effective insulin concentrations where insulin stimulation and hexose uptake were both conducted at each given assay temperature. Basal uptake was only subtly affected. Control and maximally insulin-stimulated cells incubated at 35 degrees C subsequently exhibited minimal temperature-sensitivity of uptake measured between 30 and 43 degrees C. The data are mostly consistent with the concept that insulin-sensitive glucose transporters are, after stimulation by insulin, functionally similar to basal transporters. Adipocyte plasma membranes were labelled with various spin- and fluorescence-label probes in lipid structural studies. The temperature-dependence of the order parameter S calculated from membranes labelled with 5-nitroxide stearate indicated the presence of a lipid phase change at approx. 33 degrees C. Membranes labelled with the fluorescence label 1,6-diphenylhexa-1,3,5-triene, or the cholesterol-like spin label nitroxide cholestane, reveal sharp transitions at lower temperatures. We suggest that a thermotropic lipid phase separation occurs in the adipocyte membrane that may be correlated with the temperature-dependence of hexose transport and insulin action in the intact cells. PMID:6324752

Oxygen-sensitive regulatory volume increase and Na transport in red blood cells from the cane toad, Bufo marinus.

PubMed

Kristensen, Karina; Koldkjaer, Pia; Berenbrink, Michael; Wang, Tobias

2007-07-01

The red blood cells (RBCs) of cane toad, Bufo marinus, are only partially saturated with oxygen in most of the circulation due to cardiac shunts that cause desaturation of arterial blood. The present study examines the oxygen dependency of RBC ouabain-insensitive unidirectional Na transport, using 22Na, in control cells and in cells exposed to hyperosmotic shrinkage or the beta-adrenergic agonist isoproterenol. Deoxygenation per se induced a slow, but significant Na influx, which was paralleled by a slow increase in RBC volume. Hyperosmotic shrinkage by a calculated 25% activated a robust Na influx that in the first 30 min had a strong PO2 dependency with maximal activation at low PO2 values and a P50 of approximately 5.5 kPa. This activation was completely abolished by the Na/H exchanger (NHE) inhibitor EIPA (10(-4) mol l(-1)). Hyperosmotic shrinkage is particularly interesting in B. marinus as it withstands considerable elevation in extracellular osmolarity following dehydration. Parallel studies showed that deoxygenated B. marinus RBCs had a much faster regulatory volume increase (RVI) response than air-equilibrated RBCs, reflecting the difference in magnitude of Na influxes at the two PO2 values. The extent of RVI ( approximately 60%) after 90 min, however, was similar under the two conditions, reflecting a more prolonged elevation of the shrinkage-induced Na influx in air-equilibrated RBCs. There were no significant differences in the ability to perform RVI between whole blood cells at a PCO2 of 1 and 3 kPa or washed RBCs, and 10(-4) mol l(-1) amiloride reduced the RVI under all conditions, whereas 10(-5) mol l(-1) bumetanide had no effect. Isoproterenol (10(-5) mol l(-1)) induced a significant and prolonged increase in an EIPA-sensitive and bumetanide-insensitive Na influx at low PO2 under iso-osmotic conditions, whilst there was no stimulation by isoproterenol for up to 45 min in air-equilibrated RBCs. The prolonged beta-adrenergic activation of the Na

Brain Na+, K+-ATPase Activity In Aging and Disease

PubMed Central

de Lores Arnaiz, Georgina Rodríguez; Ordieres, María Graciela López

2014-01-01

Na+/K+ pump or sodium- and potassium-activated adenosine 5’-triphosphatase (Na+, K+-ATPase), its enzymatic version, is a crucial protein responsible for the electrochemical gradient across the cell membranes. It is an ion transporter, which in addition to exchange cations, is the ligand for cardenolides. This enzyme regulates the entry of K+ with the exit of Na+ from cells, being the responsible for Na+/K+ equilibrium maintenance through neuronal membranes. This transport system couples the hydrolysis of one molecule of ATP to exchange three sodium ions for two potassium ions, thus maintaining the normal gradient of these cations in animal cells. Oxidative metabolism is very active in brain, where large amounts of chemical energy as ATP molecules are consumed, mostly required for the maintenance of the ionic gradients that underlie resting and action potentials which are involved in nerve impulse propagation, neurotransmitter release and cation homeostasis. Protein phosphorylation is a key process in biological regulation. At nervous system level, protein phosphorylation is the major molecular mechanism through which the function of neural proteins is modulted in response to extracellular signals, including the response to neurotransmitter stimuli. It is the major mechanism of neural plasticity, including memory processing. The phosphorylation of Na+, K+-ATPase catalytic subunit inhibits enzyme activity whereas the inhibition of protein kinase C restores the enzyme activity. The dephosphorylation of neuronal Na+, K+-ATPase is mediated by calcineurin, a serine / threonine phosphatase. The latter enzyme is involved in a wide range of cellular responses to Ca2+ mobilizing signals, in the regulation of neuronal excitability by controlling the activity of ion channels, in the release of neurotransmitters and hormones, as well as in synaptic plasticity and gene transcription. In the present article evidence showing Na+, K+-ATPase involvement in signaling pathways

High-Resolution Structure and Mechanism of an F/V-Hybrid Rotor Ring in a Na+-coupled ATP Synthase

PubMed Central

Matthies, Doreen; Zhou, Wenchang; Klyszejko, Adriana L.; Anselmi, Claudio; Yildiz, Özkan; Brandt, Karsten; Müller, Volker; Faraldo-Gómez, José D.; Meier, Thomas

2014-01-01

All rotary ATPases catalyze the interconversion of ATP and ADP-Pi through a mechanism that is coupled to the transmembrane flow of H+ or Na+. Physiologically, however, F/A-type enzymes specialize in ATP synthesis driven by downhill ion diffusion, while eukaryotic V-type ATPases function as ion pumps. To begin to rationalize the molecular basis for this functional differentiation, we solved the crystal structure of the Na+-driven membrane rotor of the Acetobacterium woodii ATP synthase, at 2.1 Å resolution. Unlike known structures, this rotor ring is a 9:1 heteromer of F- and V-type c-subunits, and therefore features a hybrid configuration of ion-binding sites along its circumference. Molecular and kinetic simulations are used to dissect the mechanisms of Na+ recognition and rotation of this c-ring, and to explain the functional implications of the V-type c-subunit. These structural and mechanistic insights indicate an evolutionary path between synthases and pumps involving adaptations in the rotor ring. PMID:25381992

Numerical simulation of in-situ chemical oxidation (ISCO) and biodegradation of petroleum hydrocarbons using a coupled model for bio-geochemical reactive transport

NASA Astrophysics Data System (ADS)

Marin, I. S.; Molson, J. W.

2013-05-01

Petroleum hydrocarbons (PHCs) are a major source of groundwater contamination, being a worldwide and well-known problem. Formed by a complex mixture of hundreds of organic compounds (including BTEX - benzene, toluene, ethylbenzene and xylenes), many of which are toxic and persistent in the subsurface and are capable of creating a serious risk to human health. Several remediation technologies can be used to clean-up PHC contamination. In-situ chemical oxidation (ISCO) and intrinsic bioremediation (IBR) are two promising techniques that can be applied in this case. However, the interaction of these processes with the background aquifer geochemistry and the design of an efficient treatment presents a challenge. Here we show the development and application of BIONAPL/Phreeqc, a modeling tool capable of simulating groundwater flow, contaminant transport with coupled biological and geochemical processes in porous or fractured porous media. BIONAPL/Phreeqc is based on the well-tested BIONAPL/3D model, using a powerful finite element simulation engine, capable of simulating non-aqueous phase liquid (NAPL) dissolution, density-dependent advective-dispersive transport, and solving the geochemical and kinetic processes with the library Phreeqc. To validate the model, we compared BIONAPL/Phreeqc with results from the literature for different biodegradation processes and different geometries, with good agreement. We then used the model to simulate the behavior of sodium persulfate (NaS2O8) as an oxidant for BTEX degradation, coupled with sequential biodegradation in a 2D case and to evaluate the effect of inorganic geochemistry reactions. The results show the advantages of a treatment train remediation scheme based on ISCO and IBR. The numerical performance and stability of the integrated BIONAPL/Phreeqc model was also verified.

Numerical Modeling of Coupled Water Flow and Heat Transport in Soil and Snow

NASA Astrophysics Data System (ADS)

Kelleners, T.

2015-12-01

A numerical model is developed to calculate coupled water flow and heat transport in seasonally frozen soil and snow. Both liquid water flow and water vapor flow are included. The effect of dissolved ions on soil water freezing point depression is included by combining an expression for osmotic head with the Clapeyron equation and the van Genuchten soil water retention function. The coupled water flow and heat transport equations are solved using the Thomas algorithm and Picard iteration. Ice pressure is always assumed zero and frost heave is neglected. The new model is tested using data from a high-elevation rangeland soil that is subject to significant soil freezing and a mountainous forest soil that is snow-covered for about 8 months of the year. Soil hydraulic parameters are mostly based on measurements and only vegetation parameters are fine-tuned to match measured and calculated soil water content, soil & snow temperature, and snow height. Modeling statistics for both systems show good performance for temperature, intermediate performance for snow height, and relatively low performance for soil water content, in accordance with earlier results with an older version of the model.

Mass-corrections for the conservative coupling of flow and transport on collocated meshes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Waluga, Christian, E-mail: waluga@ma.tum.de; Wohlmuth, Barbara; Rüde, Ulrich

2016-01-15

Buoyancy-driven flow models demand a careful treatment of the mass-balance equation to avoid spurious source and sink terms in the non-linear coupling between flow and transport. In the context of finite-elements, it is therefore commonly proposed to employ sufficiently rich pressure spaces, containing piecewise constant shape functions to obtain local or even strong mass-conservation. In three-dimensional computations, this usually requires nonconforming approaches, special meshes or higher order velocities, which make these schemes prohibitively expensive for some applications and complicate the implementation into legacy code. In this paper, we therefore propose a lean and conservatively coupled scheme based on standard stabilizedmore » linear equal-order finite elements for the Stokes part and vertex-centered finite volumes for the energy equation. We show that in a weak mass-balance it is possible to recover exact conservation properties by a local flux-correction which can be computed efficiently on the control volume boundaries of the transport mesh. We discuss implementation aspects and demonstrate the effectiveness of the flux-correction by different two- and three-dimensional examples which are motivated by geophysical applications.« less

Spin transport study in a Rashba spin-orbit coupling system

PubMed Central

Mei, Fuhong; Zhang, Shan; Tang, Ning; Duan, Junxi; Xu, Fujun; Chen, Yonghai; Ge, Weikun; Shen, Bo

2014-01-01

One of the most important topics in spintronics is spin transport. In this work, spin transport properties of two-dimensional electron gas in AlxGa1-xN/GaN heterostructure were studied by helicity-dependent photocurrent measurements at room temperature. Spin-related photocurrent was detected under normal incidence of a circularly polarized laser with a Gaussian distribution. On one hand, spin polarized electrons excited by the laser generate a diffusive spin polarization current, which leads to a vortex charge current as a result of anomalous circular photogalvanic effect. On the other hand, photo-induced spin polarized electrons driven by a longitudinal electric field give rise to a transverse current via anomalous Hall Effect. Both of these effects originated from the Rashba spin-orbit coupling. By analyzing spin-related photocurrent varied with laser position, the contributions of the two effects were differentiated and the ratio of the spin diffusion coefficient to photo-induced anomalous spin Hall mobility Ds/μs = 0.08 V was extracted at room temperature. PMID:24504193

ABA and GA3 increase carbon allocation in different organs of grapevine plants by inducing accumulation of non-structural carbohydrates in leaves, enhancement of phloem area and expression of sugar transporters.

PubMed

Murcia, Germán; Pontin, Mariela; Reinoso, Herminda; Baraldi, Rita; Bertazza, Gianpaolo; Gómez-Talquenca, Sebastián; Bottini, Rubén; Piccoli, Patricia N

2016-03-01

Grape quality for winemaking depends on sugar accumulation and metabolism in berries. Abscisic acid (ABA) and gibberellins (GAs) have been reported to control sugar allocation in economically important crops, although the mechanisms involved are still unknown. The present study tested if ABA and gibberellin A3 (GA3) enhance carbon allocation in fruits of grapevines by modifying phloem loading, phloem area and expression of sugar transporters in leaves and berries. Pot-grown Vitis vinifera cv. Malbec plants were sprayed with ABA and GA3 solutions. The amount of soluble sugars in leaves and berries related to photosynthesis were examined at three points of berry growth: pre-veraison, full veraison and post-veraison. Starch levels and amylase activity in leaves, gene expression of sugar transporters in leaves and berries and phloem anatomy were examined at full veraison. Accumulation of glucose and fructose in berries was hastened in ABA-treated plants at the stage of full veraison, which was correlated with enhancement of Vitis vinifera HEXOSE TRANSPORTER 2 (VvHT2) and Vitis vinifera HEXOSE TRANSPORTER 6 (VvHT6) gene expression, increases of phloem area and sucrose content in leaves. On the other hand, GA3 increased the quantity of photoassimilates delivered to the stem thus increasing xylem growth. In conclusion, stimulation of sugar transport by ABA and GA3 to berries and stems, respectively, was due to build-up of non-structural carbohydrates in leaves, modifications in phloem tissue and modulation in gene expression of sugar transporters. © 2015 Scandinavian Plant Physiology Society.

Human SLC4A11 Is a Novel NH3/H+ Co-transporter*

PubMed Central

Zhang, Wenlin; Ogando, Diego G.; Bonanno, Joseph A.; Obukhov, Alexander G.

2015-01-01

SLC4A11 has been proposed to be an electrogenic membrane transporter, permeable to Na+, H+ (OH−), bicarbonate, borate, and NH4+. Recent studies indicate, however, that neither bicarbonate or borate is a substrate. Here, we examined potential NH4+, Na+, and H+ contributions to electrogenic ion transport through SLC4A11 stably expressed in Na+/H+ exchanger-deficient PS120 fibroblasts. Inward currents observed during exposure to NH4Cl were determined by the [NH3]o, not [NH4+]o, and current amplitudes varied with the [H+] gradient. These currents were relatively unaffected by removal of Na+, K+, or Cl− from the bath but could be reduced by inclusion of NH4Cl in the pipette solution. Bath pH changes alone did not generate significant currents through SLC4A11, except immediately following exposure to NH4Cl. Reversal potential shifts in response to changing [NH3]o and pHo suggested an NH3/H+-coupled transport mode for SLC4A11. Proton flux through SLC4A11 in the absence of ammonia was relatively small, suggesting that ammonia transport is of more physiological relevance. Methylammonia produced currents similar to NH3 but with reduced amplitude. Estimated stoichiometry of SLC4A11 transport was 1:2 (NH3/H+). NH3-dependent currents were insensitive to 10 μm ethyl-isopropyl amiloride or 100 μm 4,4′- diisothiocyanatostilbene-2,2′-disulfonic acid. We propose that SLC4A11 is an NH3/2H+ co-transporter exhibiting unique characteristics. PMID:26018076

Induction of the high-affinity Na(+)-dependent glutamate transport system XAG- by hypertonic stress in the renal epithelial cell line NBL-1.

PubMed Central

Ferrer-Martinez, A; Felipe, A; Nicholson, B; Casado, J; Pastor-Anglada, M; McGivan, J

1995-01-01

The high-affinity Na(+)-dependent glutamate transport system XAG- is induced (threefold increase in Vmax. with no change in Km) by hypertonicity in the renal epithelial cell line NBL-1. This effect is dependent on protein synthesis and glycosylation and is accompanied by an increase in EAAC1 mRNA levels. Other Na(+)-dependent transport systems in this cell line do not respond to hypertonic stress. In contrast to recent findings [Ruiz-Montasell, Gomez-Angelats, Casado, Felipe, McGivan and Pastor-Anglada (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 9569-9573] showing that increased system A activity after hyperosmotic shock results from induction of a regulatory protein, this is the first demonstration that hypertonicity may increase the expression of the gene for an amino acid transport protein itself. Images Figure 4 PMID:7654212

Generalized analytical solutions to sequentially coupled multi-species advective-dispersive transport equations in a finite domain subject to an arbitrary time-dependent source boundary condition

NASA Astrophysics Data System (ADS)

Chen, Jui-Sheng; Liu, Chen-Wuing; Liang, Ching-Ping; Lai, Keng-Hsin

2012-08-01

SummaryMulti-species advective-dispersive transport equations sequentially coupled with first-order decay reactions are widely used to describe the transport and fate of the decay chain contaminants such as radionuclide, chlorinated solvents, and nitrogen. Although researchers attempted to present various types of methods for analytically solving this transport equation system, the currently available solutions are mostly limited to an infinite or a semi-infinite domain. A generalized analytical solution for the coupled multi-species transport problem in a finite domain associated with an arbitrary time-dependent source boundary is not available in the published literature. In this study, we first derive generalized analytical solutions for this transport problem in a finite domain involving arbitrary number of species subject to an arbitrary time-dependent source boundary. Subsequently, we adopt these derived generalized analytical solutions to obtain explicit analytical solutions for a special-case transport scenario involving an exponentially decaying Bateman type time-dependent source boundary. We test the derived special-case solutions against the previously published coupled 4-species transport solution and the corresponding numerical solution with coupled 10-species transport to conduct the solution verification. Finally, we compare the new analytical solutions derived for a finite domain against the published analytical solutions derived for a semi-infinite domain to illustrate the effect of the exit boundary condition on coupled multi-species transport with an exponential decaying source boundary. The results show noticeable discrepancies between the breakthrough curves of all the species in the immediate vicinity of the exit boundary obtained from the analytical solutions for a finite domain and a semi-infinite domain for the dispersion-dominated condition.

Mutations in the Na+/Citrate Cotransporter NaCT (SLC13A5) in Pediatric Patients with Epilepsy and Developmental Delay

PubMed Central

Klotz, Jenna; Porter, Brenda E; Colas, Claire; Schlessinger, Avner; Pajor, Ana M

2016-01-01

Mutations in the SLC13A5 gene that codes for the Na+/citrate cotransporter, NaCT, are associated with early onset epilepsy, developmental delay and tooth dysplasia in children. In this study, we identify additional SLC13A5 mutations in nine epilepsy patients from six families. To better characterize the syndrome, families with affected children answered questions about the scope of illness and the treatment strategies. Currently, there are no effective treatments, but some antiepileptic drugs targeting the γ-aminobutyric acid system reduce seizure frequency. Acetazolamide, a carbonic anhydrase inhibitor and atypical antiseizure medication, decreases seizures in four patients. In contrast to previous reports, the ketogenic diet and fasting resulted in worsening of symptoms. The effects of the mutations on NaCT transport function and protein expression were examined by transient transfections of COS-7 cells. There was no transport activity from any of the mutant transporters, although some of the mutant transporter proteins were present on the plasma membrane. The structural model of NaCT suggests that these mutations can affect helix packing or substrate binding. We tested various treatments, including chemical chaperones and low temperatures, but none improved transport function in the NaCT mutants. Interestingly, coexpression of NaCT and the mutants results in decreased protein expression and activity of the wild-type transporter, indicating functional interaction. In conclusion, this study has identified additional SLC13A5 mutations in patients with chronic epilepsy starting in the neonatal period, with the mutations producing inactive Na+/citrate transporters. PMID:27261973

The Conformational Changes Induced by Ubiquinone Binding in the Na+-pumping NADH:Ubiquinone Oxidoreductase (Na+-NQR) Are Kinetically Controlled by Conserved Glycines 140 and 141 of the NqrB Subunit*

PubMed Central

Strickland, Madeleine; Juárez, Oscar; Neehaul, Yashvin; Cook, Darcie A.; Barquera, Blanca; Hellwig, Petra

2014-01-01

Na+-pumping NADH:ubiquinone oxidoreductase (Na+-NQR) is responsible for maintaining a sodium gradient across the inner bacterial membrane. This respiratory enzyme, which couples sodium pumping to the electron transfer between NADH and ubiquinone, is not present in eukaryotes and as such could be a target for antibiotics. In this paper it is shown that the site of ubiquinone reduction is conformationally coupled to the NqrB subunit, which also hosts the final cofactor in the electron transport chain, riboflavin. Previous work showed that mutations in conserved NqrB glycine residues 140 and 141 affect ubiquinone reduction and the proper functioning of the sodium pump. Surprisingly, these mutants did not affect the dissociation constant of ubiquinone or its analog HQNO (2-n-heptyl-4-hydroxyquinoline N-oxide) from Na+-NQR, which indicates that these residues do not participate directly in the ubiquinone binding site but probably control its accessibility. Indeed, redox-induced difference spectroscopy showed that these mutations prevented the conformational change involved in ubiquinone binding but did not modify the signals corresponding to bound ubiquinone. Moreover, data are presented that demonstrate the NqrA subunit is able to bind ubiquinone but with a low non-catalytically relevant affinity. It is also suggested that Na+-NQR contains a single catalytic ubiquinone binding site and a second site that can bind ubiquinone but is not active. PMID:25006248

Hxt-carrier-mediated glucose efflux upon exposure of Saccharomyces cerevisiae to excess maltose.

PubMed

Jansen, Mickel L A; De Winde, Johannes H; Pronk, Jack T

2002-09-01

When wild-type Saccharomyces cerevisiae strains pregrown in maltose-limited chemostat cultures were exposed to excess maltose, release of glucose into the external medium was observed. Control experiments confirmed that glucose release was not caused by cell lysis or extracellular maltose hydrolysis. To test the hypothesis that glucose efflux involved plasma membrane glucose transporters, experiments were performed with an S. cerevisiae strain in which all members of the hexose transporter (HXT) gene family had been eliminated and with an isogenic reference strain. Glucose efflux was virtually eliminated in the hexose-transport-deficient strain. This constitutes experimental proof that Hxt transporters facilitate export of glucose from S. cerevisiae cells. After exposure of the hexose-transport-deficient strain to excess maltose, an increase in the intracellular glucose level was observed, while the concentrations of glucose 6-phosphate and ATP remained relatively low. These results demonstrate that glucose efflux can occur as a result of uncoordinated expression of the initial steps of maltose metabolism and the subsequent reactions in glucose dissimilation. This is a relevant phenomenon for selection of maltose-constitutive strains for baking and brewing.

Salt stress reduces kernel number of corn by inhibiting plasma membrane H+-ATPase activity.

PubMed

Jung, Stephan; Hütsch, Birgit W; Schubert, Sven

2017-04-01

Salt stress affects yield formation of corn (Zea mays L.) at various physiological levels resulting in an overall grain yield decrease. In this study we investigated how salt stress affects kernel development of two corn cultivars (cvs. Pioneer 3906 and Fabregas) at and shortly after pollination. In an earlier study, we found an accumulation of hexoses in the kernel tissue. Therefore, it was hypothesized that hexose uptake into developing endosperm and embryo might be inhibited. Hexoses are transported into the developing endosperm by carriers localized in the plasma membrane (PM). The transport is driven by the pH gradient which is built up by the PM H + -ATPase. It was investigated whether the PM H + -ATPase activity in developing corn kernels was inhibited by salt stress, which would cause a lower pH gradient resulting in impaired hexose import and finally in kernel abortion. Corn grown under control and salt stress conditions was harvested 0 and 2 days after pollination (DAP). Under salt stress sucrose and hexose concentrations in kernel tissue were higher 0 and 2 DAP. Kernel PM H + -ATPase activity was not affected at 0 DAP, but it was reduced at 2 DAP. This is in agreement with the finding, that kernel growth and thus kernel setting was not affected in the salt stress treatment at pollination, but it was reduced 2 days later. It is concluded that inhibition of PM H + -ATPase under salt stress impaired the energization of hexose transporters into the cells, resulting in lower kernel growth and finally in kernel abortion. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

A natural protective mechanism against hyperglycaemia in vascular endothelial and smooth-muscle cells: role of glucose and 12-hydroxyeicosatetraenoic acid.

PubMed Central

Alpert, Evgenia; Gruzman, Arie; Totary, Hanan; Kaiser, Nurit; Reich, Reuven; Sasson, Shlomo

2002-01-01

Bovine aortic endothelial and smooth-muscle cells down-regulate the rate of glucose transport in the face of hyperglycaemia, thus providing protection against deleterious effects of increased intracellular glucose levels. When exposed to high glucose concentrations these cells reduced the mRNA and protein content of their typical glucose transporter, GLUT-1, as well as its plasma-membrane abundance. Inhibition of the lipoxygenase (LO) pathway, and particularly 12-LO, reversed this glucose-induced down-regulatory process and restored the rate of hexose transport to the level seen in vascular cells exposed to normal glucose levels. This reversal was accompanied by increased levels of GLUT-1 mRNA and protein, as well as of its plasma-membrane content. Exposure of the vascular cells to elevated glucose concentrations increased by 2-3-fold the levels of cell-associated and secreted 12-hydroxyeicosatetraenoic acid (12-HETE), the product of 12-LO. Inhibition of 15- and 5-LO, cyclo-oxygenases 1 and 2, and eicosanoid-producing cytochrome P450 did not modify the hexose-transport system in vascular cells. These results suggest a role for HETEs in the autoregulation of hexose transport in vascular cells. 8-Iso prostaglandin F(2alpha), a non-enzymic oxidation product of arachidonic acid, had no effect on the hexose-transport system in vascular cells exposed to hyperglycaemic conditions. Taken together, these findings show that hyperglycaemia increases the production rate of 12-HETE, which in turn mediates the down-regulation of GLUT-1 expression and the glucose-transport system in vascular endothelial and smooth-muscle cells. PMID:11853550

Expression of apical Na(+)-L-glutamine co-transport activity, B(0)-system neutral amino acid co-transporter (B(0)AT1) and angiotensin-converting enzyme 2 along the jejunal crypt-villus axis in young pigs fed a liquid formula.

PubMed

Yang, Chengbo; Yang, Xiaojian; Lackeyram, Dale; Rideout, Todd C; Wang, Zirong; Stoll, Barbara; Yin, Yulong; Burrin, Douglas G; Fan, Ming Z

2016-06-01

Gut apical amino acid (AA) transport activity is high at birth and during suckling, thus being essential to maintain luminal nutrient-dependent mucosal growth through providing AA as essential metabolic fuel, substrates and nutrient stimuli for cellular growth. Because system-B(0) Na(+)-neutral AA co-transporter (B(0)AT1, encoded by the SLC6A19 gene) plays a dominant role for apical uptake of large neutral AA including L-Gln, we hypothesized that high apical Na(+)-Gln co-transport activity, and B(0)AT1 (SLC6A19) in co-expression with angiotensin-converting enzyme 2 (ACE2) were expressed along the entire small intestinal crypt-villus axis in young animals via unique control mechanisms. Kinetics of Na(+)-Gln co-transport activity in the apical membrane vesicles, prepared from epithelial cells sequentially isolated along the jejunal crypt-villus axis from liquid formula-fed young pigs, were measured with the membrane potential being clamped to zero using thiocyanate. Apical maximal Na(+)-Gln co-transport activity was much higher (p < 0.05) in the upper villus cells than in the middle villus (by 29 %) and the crypt (by 30 %) cells, whereas Na(+)-Gln co-transport affinity was lower (p < 0.05) in the upper villus cells than in the middle villus and the crypt cells. The B(0)AT1 (SLC6A19) mRNA abundance was lower (p < 0.05) in the crypt (by 40-47 %) than in the villus cells. There were no significant differences in B(0)AT1 and ACE2 protein abundances on the apical membrane among the upper villus, the middle villus and the crypt cells. Our study suggests that piglet fast growth is associated with very high intestinal apical Na(+)-neutral AA uptake activities via abundantly co-expressing B(0)AT1 and ACE2 proteins in the apical membrane and by transcribing the B(0)AT1 (SLC6A19) gene in the epithelia along the entire crypt-villus axis.

Mechanistic and Kinetic Analysis of Na2SO4-Modified Laterite Decomposition by Thermogravimetry Coupled with Mass Spectrometry

PubMed Central

Yang, Song; Du, Wenguang; Shi, Pengzheng; Shangguan, Ju; Liu, Shoujun; Zhou, Changhai; Chen, Peng; Zhang, Qian; Fan, Huiling

2016-01-01

Nickel laterites cannot be effectively used in physical methods because of their poor crystallinity and fine grain size. Na2SO4 is the most efficient additive for grade enrichment and Ni recovery. However, how Na2SO4 affects the selective reduction of laterite ores has not been clearly investigated. This study investigated the decomposition of laterite with and without the addition of Na2SO4 in an argon atmosphere using thermogravimetry coupled with mass spectrometry (TG-MS). Approximately 25 mg of samples with 20 wt% Na2SO4 was pyrolyzed under a 100 ml/min Ar flow at a heating rate of 10°C/min from room temperature to 1300°C. The kinetic study was based on derivative thermogravimetric (DTG) curves. The evolution of the pyrolysis gas composition was detected by mass spectrometry, and the decomposition products were analyzed by X-ray diffraction (XRD). The decomposition behavior of laterite with the addition of Na2SO4 was similar to that of pure laterite below 800°C during the first three stages. However, in the fourth stage, the dolomite decomposed at 897°C, which is approximately 200°C lower than the decomposition of pure laterite. In the last stage, the laterite decomposed and emitted SO2 in the presence of Na2SO4 with an activation energy of 91.37 kJ/mol. The decomposition of laterite with and without the addition of Na2SO4 can be described by one first-order reaction. Moreover, the use of Na2SO4 as the modification agent can reduce the activation energy of laterite decomposition; thus, the reaction rate can be accelerated, and the reaction temperature can be markedly reduced. PMID:27333072

Effects of surface functionalization of hydrophilic NaYF4 nanocrystals doped with Eu3+ on glutamate and GABA transport in brain synaptosomes

NASA Astrophysics Data System (ADS)

Sojka, Bartlomiej; Kociołek, Daria; Banski, Mateusz; Borisova, Tatiana; Pozdnyakova, Natalia; Pastukhov, Artem; Borysov, Arsenii; Dudarenko, Marina; Podhorodecki, Artur

2017-08-01

Specific rare earth doped nanocrystals (NCs), a recent class of nanoparticles with fluorescent features, have great bioanalytical potential. Neuroactive properties of NaYF4 nanocrystals doped with Eu3+ were assessed based on the analysis of their effects on glutamate- and γ-aminobutyric acid (GABA) transport process in nerve terminals isolated from rat brain (synaptosomes). Two types of hydrophilic NCs were examined in this work: (i) coated by polyethylene glycol (PEG) and (ii) with OH groups at the surface. It was found that NaYF4:Eu3+-PEG and NaYF4:Eu3+-OH within the concentration range of 0.5-3.5 and 0.5-1.5 mg/ml, respectively, did not influence Na+-dependent transporter-dependent l-[14C]glutamate and [3H]GABA uptake and the ambient level of the neurotransmitters in the synaptosomes. An increase in NaYF4:Eu3+-PEG and NaYF4:Eu3+-OH concentrations up to 7.5 and 3.5 mg/ml, respectively, led to the (1) attenuation of the initial velocity of uptake of l-[14C]glutamate and [3H]GABA and (2) elevation of ambient neurotransmitters in the suspension of nerve terminals. In the mentioned concentrations, nanocrystals did not influence acidification of synaptic vesicles that was shown with pH-sensitive fluorescent dye acridine orange, however, decreased the potential of the plasma membrane of synaptosomes. In comparison with other nanoparticles studied with similar methodological approach, NCs start to exhibit their effects on neurotransmitter transport at concentrations several times higher than those shown for carbon dots, detonation nanodiamonds and an iron storage protein ferritin, whose activity can be registered at 0.08, 0.5 and 0.08 mg/ml, respectively. Therefore, NCs can be considered lesser neurotoxic as compared to above nanoparticles.

Day–Night Changes of Energy-rich Compounds in Crassulacean Acid Metabolism (CAM) Species Utilizing Hexose and Starch

PubMed Central

CHEN, LI-SONG; NOSE, AKIHIRO

2004-01-01

• Background and Aims Plants with crassulacean acid metabolism (CAM) can be divided into two groups according to the major carbohydrates used for malic acid synthesis, either polysaccharide (starch) or monosaccharide (hexose). This is related to the mechanism and affects energy metabolism in the two groups. In Kalanchoë pinnata and K. daigremontiana, which utilize starch, ATP-dependent phosphofructokinase (tonoplast inorganic pyrophosphatase) activity is greater than inorganic pyrophosphate-dependent phosphofructokinase (tonoplast adenosine triphosphatase) activity, but the reverse is the case in pineapple (Ananas comosus) utilizing hexose. To test the hypothesis that the energy metabolism of the two groups differs, day-night changes in the contents of ATP, ADP, AMP, inorganic phosphate (Pi), phosphoenolpyruvate (PEP) and inorganic pyrophosphate (PPi) in K. pinnata and K. daigremontiana leaves and in pineapple chlorenchyma were analysed. • Methods The contents of energy-rich compounds were measured spectrophotometrically in extracts of tissue sampled in the light and dark, using potted plants, kept for 15 d before the experiments in a growth chamber. • Key Results In the three species, ATP content and adenylate energy charge (AEC) increased in the dark and decreased in the light, in contrast to ADP and AMP. Changes in ATP and AEC were greater in Kalanchoë leaves than in pineapple chlorenchyma. PPi content in the three species increased in the dark, but on illumination it decreased rapidly and substantially, remaining little changed through the rest of the light period. Pi content of Kalanchoë leaves did not change between dark and light, whereas Pi in pineapple chlorenchyma increased in the dark and decreased in the light, and the changes were far greater than in Kalanchoë leaves. Light-dark changes in PEP content in the three species were similar. • Conclusions These results corroborate our hypothesis that day–night changes in the contents of energy

Coupled effects of solution chemistry and hydrodynamics on the mobility and transport of quantum dot nanomaterials in the Vadose Zone