Amino acid transport in normal and glutathione-deficient sheep erythrocytes.
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ABSTRACT:
SUBMITTER: Young JD
PROVIDER: S-EPMC1172674 | biostudies-other | 1976 Jan
REPOSITORIES: biostudies-other
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Project description:The specificity of amino acid transport in normal (high-glutathione) sheep erythrocytes was investigated by studying the interaction of various neutral and dibasic amino acids in both competition and exchange experiments. Apparent Ki values were obtained for amino acids as inhibitors of L-alanine influx. Amino acids previously found to be transported by high-glutathione cells at fast rates (L-cysteine, L-alpha-amino-n-butyrate) were the most effective inhibitors. D-Alanine and D-alpha-amino-n-butyrate were without effect. Of the remaining amino acids studied, only L-norvaline, L-valine, L-norleucine, L-serine and L-2,4-diamino-n-butyrate significantly inhibited L-alanine uptake. L-Alanine efflux from pre-loaded cells was markedly stimulated by extracellular L-alanine. Those amino acids that inhibited L-alanine influx also stimulated L-alanine efflux. In addition, D-alanine, D-alpha-amino-n-biutyrate, L-threonine, L-asparagine, L-alpha, beta-diaminoproprionate, L-ornithine, L-lysine and S-2-aminoethyl-L-cysteine also significantly stimulated L-alanine efflux. L-Lysine uptake was inhibited by L-alanine but not by D-alanine, and the inhibitory potency of L-alanine was not influenced by the replacement of Na+ in the incubation medium with choline. L-Lysine efflux from pre-loaded cells was stimulated by L-alanine but not by D-alanine. It is concluded that these cells possess a highly selective stero-specific amino acid-transport system. Although the optimum substrates are small neutral amino acids, this system also has a significant affinity for dibasic amino acids.
| S-EPMC1164566 | biostudies-other
Project description:Resealed ghosts from pigeon erythrocytes were shown to haemolyse during incubation in isotonic media with pH values greater than about 7 and high concentrations of Na+ inside the ghosts seemed to enhance this effect. At lower pH values the ghosts were stable but still highly permeable to Na+ and K+, and moderately permeable to sucrose. Under the latter conditions the ghosts transported amino acids in a way qualitatively but not quantitatively similar to intact erythrocytes. The Na+-dependent transport of serine and alanine by the ghosts consisted essentially of an exchange of extracellular for intracellular amino acids, with no significant net flux. In contrast, net fluxes of glycine in the direction of the Na+-concentration gradient across the ghost membrane were demonstrated. However, under one condition a small net influx of glycine occurred against the prevailing Na+-concentration gradient. Unlike Na+-dependent glycine uptake, the uptake of six other amino acids by intact pigeon erythrocytes was not influenced by the nature of the anion present. The significance of these findings in relation to previous work on the Na+-gradient hypothesis of membrane transport is discussed.
| S-EPMC1158154 | biostudies-other
Project description:In many cell types, including human erythrocytes, membrane transport of hydrophobic amino acids such as leucine and phenylalanine is mediated primarily by Na(+)-independent system L. In this paper we demonstrate that erythrocytes from the rat have a 400-fold higher system L transport capacity than human erythrocytes. We have exploited this high transport activity to achieve the first successful reconstitution of an erythrocyte amino acid transporter into phospholipid vesicles. Rat erythrocyte membranes were depleted of extrinsic membrane proteins, solubilized in 50 mM n-octyl glucoside and reconstituted into egg-yolk phospholipid vesicles by a gel filtration freeze-thaw protocol. Optimal reconstitution of transport activity occurred at lipid/protein ratios of 25-35:1. At a lipid/protein ratio of 25:1, one-half of the total uptake of L-[14C]leucine (0.2 mM, 25 degrees C) was inhibited by 2 mM phloretin and thus judged to be carrier-mediated. This component of L-leucine uptake was inhibited by non-radioactive L-phenylalanine and L-leucine, and only to a very much weaker extent by glycine and L-alanine. Two other inhibitors of system L in intact cells, MK196 and PCMBS (p-chloromercuriphenylsulphonate), were also effective inhibitors of phloretin-sensitive L-leucine transport in reconstituted proteoliposomes. Phloretin-insensitive uptake of L-leucine in proteoliposomes occurred by simple diffusion across the lipid bilayer.
| S-EPMC1134163 | biostudies-other
Project description:BackgroundCitrin (mitochondrial aspartate-glutamate transporter) deficiency causes the failures in both carbohydrate-energy metabolism and the urea cycle, and the alterations in the serum levels of several amino acids in the stages of newborn (NICCD) and adult (CTLN2). However, the clinical manifestations are resolved between the NICCD and CTLN2, but the reasons are still unclear. This study evaluated the serum amino acid profile in citrin-deficient children during the healthy stage.MethodsUsing HPLC-MS/MS analysis, serum amino acids were evaluated among 20 citrin-deficient children aged 5-13 years exhibiting normal liver function and 35 age-matched healthy controls.ResultsThe alterations in serum amino acids characterized in the NICCD and CTLN2 stages were not observed in the citrin-deficient children. Amino acids involved in the urea cycle, including arginine, ornithine, citrulline, and aspartate, were comparable in the citrin-deficient children to the respective control levels, but serum urea was twofold higher, suggestive of a functional urea cycle. The blood sugar level was normal, but glucogenic amino acids and glutamine were significantly decreased in the citrin-deficient children compared to those in the controls. In addition, significant increases of ketogenic amino acids, branched-chain amino acids (BCAAs), a valine intermediate 3-hydroxyisobutyrate, and ?-alanine were also found in the citrin-deficient children.ConclusionThe profile of serum amino acids in the citrin-deficient children during the healthy stage showed different characteristics from the NICCD and CTLN2 stages, suggesting that the failures in both urea cycle function and energy metabolism might be compensated by amino acid metabolism.SynopsisIn the citrin-deficient children during the healthy stage, the characteristics of serum amino acids, including decrease of glucogenic amino acids, and increase of ketogenic amino acids, BCAAs, valine intermediate, and ?-alanine, were found by comparison to the age-matched healthy control children, and it suggested that the characteristic alteration of serum amino acids may be resulted from compensation for energy metabolism and ammonia detoxification.
| S-EPMC6323014 | biostudies-literature
Project description:Protein supplementation during human pregnancy does not improve fetal growth and may increase small-for-gestational-age birth rates and mortality. To define possible mechanisms, sheep with twin pregnancies were infused with amino acids (AA group, n = 7) or saline (C group, n = 4) for 4 days during late gestation. In the AA group, fetal plasma leucine, isoleucine, valine, and lysine concentrations were increased (P < 0.05), and threonine was decreased (P < 0.05). In the AA group, fetal arterial pH (7.365 +/- 0.007 day 0 vs. 7.336 +/- 0.012 day 4, P < 0.005), hemoglobin-oxygen saturation (46.2 +/- 2.6 vs. 37.8 +/- 3.6%, P < 0.005), and total oxygen content (3.17 +/- 0.17 vs. 2.49 +/- 0.20 mmol/l, P < 0.0001) were decreased on day 4 compared with day 0. Fetal leucine disposal did not change (9.22 +/- 0.73 vs. 8.09 +/- 0.63 micromol x min(-1) x kg(-1), AA vs. C), but the rate of leucine oxidation increased 43% in the AA group (2.63 +/- 0.16 vs. 1.84 +/- 0.24 micromol x min(-1) x kg(-1), P < 0.05). Fetal oxygen utilization tended to be increased in the AA group (327 +/- 23 vs. 250 +/- 29 micromol x min(-1) x kg(-1), P = 0.06). Rates of leucine incorporation into fetal protein (5.19 +/- 0.97 vs. 5.47 +/- 0.89 micromol x min(-1) x kg(-1), AA vs. C), release from protein breakdown (4.20 +/- 0.95 vs. 4.62 +/- 0.74 micromol x min(-1) x kg(-1)), and protein accretion (1.00 +/- 0.30 vs. 0.85 +/- 0.25 micromol x min(-1) x kg(-1)) did not change. Consistent with these data, there was no change in the fetal skeletal muscle ubiquitin ligases MaFBx1 or MuRF1 or in the protein synthesis regulators 4E-BP1, eEF2, eIF2alpha, and p70(S6K). Decreased concentrations of certain essential amino acids, increased amino acid oxidation, fetal acidosis, and fetal hypoxia are possible mechanisms to explain fetal toxicity during maternal amino acid supplementation.
| S-EPMC2739698 | biostudies-literature
Project description:We performed RNA-seq to investigate gene expression changes in pancreatic cancer cells following gemcitabine with high-dose L-Gln.
2022-09-30 | GSE214325 | GEO
Project description:We review the mechanisms responsible for amino acid homeostasis in Saccharomyces cerevisiae and other fungi. Amino acid homeostasis is essential for cell growth and survival. Hence, the de novo synthesis reactions, metabolic conversions, and transport of amino acids are tightly regulated. Regulation varies from nitrogen pool sensing to control by individual amino acids and takes place at the gene (transcription), protein (posttranslational modification and allostery), and vesicle (trafficking and endocytosis) levels. The pools of amino acids are controlled via import, export, and compartmentalization. In yeast, the majority of the amino acid transporters belong to the APC (amino acid-polyamine-organocation) superfamily, and the proteins couple the uphill transport of amino acids to the electrochemical proton gradient. Although high-resolution structures of yeast amino acid transporters are not available, homology models have been successfully exploited to determine and engineer the catalytic and regulatory functions of the proteins. This has led to a further understanding of the underlying mechanisms of amino acid sensing and subsequent downregulation of transport. Advances in optical microscopy have revealed a new level of regulation of yeast amino acid transporters, which involves membrane domain partitioning. The significance and the interrelationships of the latest discoveries on amino acid homeostasis are put in context.
| S-EPMC7405077 | biostudies-literature
Project description:BACKGROUND:Although reduced glutathione (rGSH) is decreased in obese mice and humans, block of GSH synthesis by buthionine sulfoximine (BSO) results in a lean, insulin-sensitive phenotype. Data is lacking about the effect of BSO on GSH precursors, cysteine and glutamate. Plasma total cysteine (tCys) is positively associated with stearoyl-coenzyme A desaturase (SCD) activity and adiposity in humans and animal models. OBJECTIVE:To explore the phenotype, amino acid and fatty acid profiles in BSO-treated mice. DESIGN:Male C3H/HeH mice aged 11 weeks were fed a high-fat diet with or without BSO in drinking water (30 mmol/L) for 8 weeks. Amino acid and fatty acid changes were assessed, as well as food consumption, energy expenditure, locomotor activity, body composition and liver vacuolation (steatosis). RESULTS:Despite higher food intake, BSO decreased particularly fat mass but also lean mass (both P<0.001), and prevented fatty liver vacuolation. Physical activity increased during the dark phase. BSO decreased plasma free fatty acids and enhanced insulin sensitivity. BSO did not alter liver rGSH, but decreased plasma total GSH (tGSH) and rGSH (by ~70%), and liver tGSH (by 82%). Glutamate accumulated in plasma and liver. Urine excretion of cysteine and its precursors was increased by BSO. tCys, rCys and cystine decreased in plasma (by 23-45%, P<0.001 for all), but were maintained in liver, at the expense of decreased taurine. Free and total plasma concentrations of the SCD products, oleic and palmitoleic acids were decreased (by 27-38%, P <0.001 for all). CONCLUSION:Counterintuitively, block of GSH synthesis decreases circulating tCys, raising the question of whether the BSO-induced obesity-resistance is linked to cysteine depletion. Cysteine-supplementation of BSO-treated mice is warranted to dissect the effects of cysteine and GSH depletion on energy metabolism.
| S-EPMC5082875 | biostudies-literature
Project description:Glioblastoma multiforme (GBM) is a deadly brain tumor with a large unmet therapeutic need. Here, we tested the hypothesis that wild-type p53 is a negative transcriptional regulator of SLC7A11, the gene encoding the System xc- (SXC) catalytic subunit, xCT, in GBM. We demonstrate that xCT expression is inversely correlated with p53 expression in patient tissue. Using representative patient derived (PDX) tumor xenolines with wild-type, null, and mutant p53 we show that p53 expression negatively correlates with xCT expression. Using chromatin immunoprecipitation studies, we present a molecular interaction whereby p53 binds to the SLC7A11 promoter, suppressing gene expression in PDX GBM cells. Accordingly, genetic knockdown of p53 increases SLC7A11 transcript levels; conversely, over-expressing p53 in p53-null GBM cells downregulates xCT expression and glutamate release. Proof of principal studies in mice with flank gliomas demonstrate that daily treatment with the mutant p53 reactivator, PRIMA-1Met, results in reduced tumor growth associated with reduced xCT expression. These findings suggest that p53 is a molecular switch for GBM glutamate biology, with potential therapeutic utility.
| S-EPMC8699180 | biostudies-literature
Project description:Amino acid transporters play very important roles in nutrient uptake, neurotransmitter recycling, protein synthesis, gene expression, cell redox balance, cell signaling, and regulation of cell volume. With regard to transporters that are closely connected to metabolism, amino acid transporter-associated diseases are linked to metabolic disorders, particularly when they involve different organs, cell types, or cell compartments. To date, 65 different human solute carrier (SLC) families and more than 400 transporter genes have been identified, including 11 that are known to include amino acid transporters. This review intends to summarize and update all the conditions in which a strong association has been found between an amino acid transporter and an inherited metabolic disorder. Many of these inherited disorders have been identified in recent years. In this work, the physiological functions of amino acid transporters will be described by the inherited diseases that arise from transporter impairment. The pathogenesis, clinical phenotype, laboratory findings, diagnosis, genetics, and treatment of these disorders are also briefly described. Appropriate clinical and diagnostic characterization of the underlying molecular defect may give patients the opportunity to avail themselves of appropriate therapeutic options in the future.
| S-EPMC6981491 | biostudies-literature