scholarly journals An explanation of the asymmetric binding of sugars to the human erythrocyte sugar-transport systems

1973 ◽  
Vol 135 (3) ◽  
pp. 539-541 ◽  
Author(s):  
J. E. G. Barnett ◽  
G. D. Holman ◽  
K. A. Munday
Keyword(s):  
Transport System ◽  
Human Erythrocyte ◽  
Sugar Transport ◽  
Transport Systems

6-O-Alkyl-d-galactoses competitively inhibit the erythrocyte sugar-transport system when added to the outside of the cells, but not to the inside. n-Propyl β-d-glucopyranoside competitively inhibits the system on the inside of the cells, but not on the outside. A model for sugar transport is proposed.

scholarly journals Evidence for two asymmetric conformational states in the human erythrocyte sugar-transport system

1975 ◽  
Vol 145 (3) ◽  
pp. 417-429 ◽  
Author(s):  
J E Barnett ◽  
G D Holman ◽  
R A Chalkley ◽  
K A Munday
Keyword(s):  
Glucose Transport ◽  
Transport System ◽  
Human Erythrocyte ◽  
Partition Coefficients ◽  
External Medium ◽  
Sugar Transport ◽  
Conformational States ◽  
Glucose Transport System ◽  
Oil Water

6-O-methyl-, 6-O-propyl-, 6-O-pentyl- and 6-O-benzyl-D-galactose, and 6-O-methyl-, 6-O-propyl- and 6-O-pentyl-D-glucose inhibit the glucose-transport system of the human erythrocyte when added to the external medium. Penetration of 6-O-methyl-D-galactose is inhibited by D-glucose, suggesting that it is transported by the glucose-transport system, but the longer-chain 6-O-alkyl-D-galactoses penetrate by a slower D-glucose-insensitive route at rates proportional to their olive oil/water partition coefficients. 6-O-n-Propyl-D-glucose and 6-O-n-propyl-D-galactose do not significantly inhibit L-sorbose entry or D-glucose exit when present only on the inside of the cells whereas propyl-beta-D-glucopyranoside, which also penetrates the membrane slowly by a glucose-insensitive route, only inhibits L-sorbose entry or D-glucose exit when present inside the cells, and not when on the outside. The 6-O-alkyl-D-galactoses, like the other nontransported C-4 and C-6 derivatives, maltose and 4,6-O-ethylidene-D-glucose, protect against fluorodinitrobenzene inactivation, whereas propyl beta-D-glucopyranoside stimulates the inactivation. Of the transported sugars tested, those modified at C-1, C-2 and C-3 enhance fluorodinitrobenzene inactivation, where those modified at C-4 and C-6 do not, but are inert or protect against inactivation. An asymmetric mechanism is proposed with two conformational states in which the sugar binds to the transport system so that C-4 and C-6 are in contact with the solvent on the outside and C-1 is in contact with the solvent on the inside of the cell. It is suggested that fluorodinitrobenzene reacts with the form of the transport system that binds sugars at the inner side of the membrane. An Appendix describes the theoretical basis of the experimental methods used for the determination of kinetic constants for non-permeating inhibitors.

scholarly journals The Glucuronic Acid Utilization Gene Cluster from Bacillus stearothermophilus T-6

1999 ◽  
Vol 181 (12) ◽  
pp. 3695-3704 ◽  
Author(s):  
Smadar Shulami ◽  
Orit Gat ◽  
Abraham L. Sonenshein ◽  
Yuval Shoham
Keyword(s):  
Gene Cluster ◽  
Transport System ◽  
Bacillus Stearothermophilus ◽  
Glucuronic Acid ◽  
Genomic Library ◽  
Regulatory Gene ◽  
Sugar Transport ◽  
Transcriptional Unit ◽  
Transport Systems ◽  
Potential Operator

ABSTRACT A λ-EMBL3 genomic library of Bacillus stearothermophilus T-6 was screened for hemicellulolytic activities, and five independent clones exhibiting β-xylosidase activity were isolated. The clones overlap each other and together represent a 23.5-kb chromosomal segment. The segment contains a cluster of xylan utilization genes, which are organized in at least three transcriptional units. These include the gene for the extracellular xylanase, xylanase T-6; part of an operon coding for an intracellular xylanase and a β-xylosidase; and a putative 15.5-kb-long transcriptional unit, consisting of 12 genes involved in the utilization of α-d-glucuronic acid (GlcUA). The first four genes in the potential GlcUA operon (orf1, -2, -3, and -4) code for a putative sugar transport system with characteristic components of the binding-protein-dependent transport systems. The most likely natural substrate for this transport system is aldotetraouronic acid [2-O-α-(4-O-methyl-α-d-glucuronosyl)-xylotriose] (MeGlcUAXyl3). The following two genes code for an intracellular α-glucuronidase (aguA) and a β-xylosidase (xynB). Five more genes (kdgK,kdgA, uxaC, uxuA, anduxuB) encode proteins that are homologous to enzymes involved in galacturonate and glucuronate catabolism. The gene cluster also includes a potential regulatory gene, uxuR, the product of which resembles repressors of the GntR family. The apparent transcriptional start point of the cluster was determined by primer extension analysis and is located 349 bp from the initial ATG codon. The potential operator site is a perfect 12-bp inverted repeat located downstream from the promoter between nucleotides +170 and +181. Gel retardation assays indicated that UxuR binds specifically to this sequence and that this binding is efficiently prevented in vitro by MeGlcUAXyl3, the most likely molecular inducer.

Effects of phloretin and theophylline on 3-O-methylglucose transport by intestinal epithelial cells

1978 ◽  
Vol 234 (3) ◽  
pp. C64-C72 ◽  
Author(s):  
J. Randles ◽  
G. A. Kimmich
Keyword(s):  
Epithelial Cells ◽  
Transport System ◽  
Intestinal Epithelial Cells ◽  
Sugar Transport ◽  
Inhibitory Effect ◽  
Metabolic Effects ◽  
Transport Systems ◽  
Facilitated Diffusion ◽  
Intestinal Epithelial ◽  
Gradient Enhancement

Phloretin and theophylline each exert an immediate inhibitory effect on the Na+-independent, facilitated-diffusion transport system for sugar associated with intestinal epithelial cells. Phloretin inhibits approximately 50% more of the total Na+-independent sugar flux than theophylline. Neither agent has an immediate effect on the Na+-dependent, concentrative sugar transport system, although preincubation of the cells with phloretin causes a significant inhibition. The slowly developing effect is correlated with a decrease in cellular adenosine triphosphate (ATP) and an elevation of intracellular Na+. Other agents which elevate cell Na+ also inhibit Na+-dependent sugar influx, even if ATP levels are not depleted. On the other hand, if ATP is depleted by phloretin under conditions in which the cells do not gain Na+, the inhibitory effect on Na+-dependent sugar flux tends to disappear. The slow-onset phloretin effects are due to transinhibition of the Na+-dependent sugar carrier by cellular Na+. When the passive sugar carrier is inhibited by phloretin or theophylline, the concentrative system can establish an enhanced sugar gradient. Because of the secondary metabolic effects of phloretin, theophylline induces a greater gradient enhancement despite its more limited effect on the passive sugar-transport system. Sugar gradients as large as 20-fold are induced by theophylline, in contrast to 12-fold gradients observed in the presence of phloretin and approximately 7- to 8-fold for untreated cells. These results are discussed in terms of conceptual questions regarding the energetics of Na+-dependent transport systems.

scholarly journals Structural requirements for binding to the sugar-transport system of the human erythrocyte

1973 ◽  
Vol 131 (2) ◽  
pp. 211-221 ◽  
Author(s):  
J. E. G. Barnett ◽  
G. D. Holman ◽  
K. A. Munday
Keyword(s):  
Transport System ◽  
Human Erythrocyte ◽  
Sugar Transport ◽  
Hydroxyl Group ◽  
Structural Requirements ◽  
Glucose Molecule ◽  
Inhibition Constants ◽  
Polar Substituents ◽  
Sugar Derivatives ◽  
Potential Substrate

The structural requirements for binding to the glucose/sorbose-transport system in the human erythrocyte were explored by measuring the inhibition constants, Ki, for specifically substituted analogues of d-glucose when l-sorbose was the penetrating sugar. Derivatives in which a hydroxyl group in the d-gluco configuration was inverted, or replaced by a hydrogen atom, at C-1, C-2, C-3, C-4 or C-6 of the d-glucose molecule, all bound to the carrier, confirming that no single hydroxyl group is essential for binding to the carrier. The binding and transport of 1-deoxy-d-glucose confirmed that the sugars bind in the pyranose form. The relative inhibition constants of d-glucose and its deoxy, epimeric and fluorinated analogues are consistent with the combination of β-d-glucopyranose with the carrier by hydrogen bonds at C-1, C-3, probably C-4, and possibly C-6 of the sugar. Both polar and non-polar substituents at C-6 enhance the affinity of d-glucose derivatives relative to d-xylose, and d-galactose derivatives relative to l-arabinose, and it is suggested that the carrier region around C-6 of the sugar may contain both hydrophobic and polar binding groups. The spatial requirements at C-1, C-2, C-3, C-4 and C-6 were explored by comparing the relative binding of d-glucose and its halogeno and O-alkyl substituents. The carrier protein closely approaches the sugar except at C-3 in the d-gluco configuration, C-4 and C-6. d-Glucal was a good inhibitor, showing that a strict chair form is not essential for binding. 3-O-(2′,3′-Epoxypropyl)-d-glucose, a potential substrate-directed alkylating agent, bound to the carrier, but did not inactivate it.

SMIT2 mediates all myo-inositol uptake in apical membranes of rat small intestine

2007 ◽  
Vol 293 (6) ◽  
pp. G1300-G1307 ◽  
Author(s):  
Rym Aouameur ◽  
Sandra Da Cal ◽  
Pierre Bissonnette ◽  
Michael J. Coady ◽  
Jean-Yves Lapointe
Keyword(s):  
Small Intestine ◽  
Transport System ◽  
Sugar Transport ◽  
Transport Systems ◽  
Affinity Constant ◽  
Rat Intestine ◽  
Electrophysiological Studies ◽  
Rabbit Intestine ◽  
Apical Membranes ◽  
Inhibition Studies

This study presents the characterization of myo-inositol (MI) uptake in rat intestine as evaluated by use of purified membrane preparations. Three secondary active MI cotransporters have been identified; two are Na+ coupled (SMIT1 and SMIT2) and one is H+ coupled (HMIT). Through inhibition studies using selective substrates such as d-chiro-inositol (DCI, specific for SMIT2) and l-fucose (specific for SMIT1), we show that SMIT2 is exclusively responsible for apical MI transport in rat intestine; rabbit intestine appears to lack apical transport of MI. Other sugar transport systems known to be present in apical membranes, such as SGLT1 or GLUT5, lacked any significant contribution to MI uptake. Functional analysis of rat SMIT2 activity, via electrophysiological studies in Xenopus oocytes, demonstrated similarities to the activities of SMIT2 from other species (rabbit and human) displaying high affinities for MI (0.150 ± 0.040 mM), DCI (0.31 ± 0.06 mM), and phlorizin (Pz; 0.016 ± 0.007 mM); low affinity for glucose (36 ± 7 mM); and no affinity for l-fucose. Although these functional characteristics essentially confirmed those found in rat intestinal apical membranes, a unique discrepancy was seen between the two systems studied in that the affinity constant for glucose was ∼40-fold lower in vesicles ( Ki = 0.94 ± 0.35 mM) than in oocytes. Finally, the transport system responsible for the basolateral efflux transporter of glucose in intestine, GLUT2, did not mediate any significant radiolabeled MI uptake in oocytes, indicating that this transport system does not participate in the basolateral exit of MI from small intestine.

Renal sugar transport in the winter flounder. III. Two glucose transport systems

1977 ◽  
Vol 232 (3) ◽  
pp. F227-F234 ◽  
Author(s):  
A. Kleinzeller ◽  
G. R. Dubyak ◽  
P. M. Griffin ◽  
E. M. McAvoy ◽  
J. M. Mullin ◽  
...  
Keyword(s):  
Hydrogen Bonding ◽  
Glucose Transport ◽  
Transport System ◽  
Sugar Transport ◽  
Winter Flounder ◽  
Transport Systems ◽  
Renal Tubules ◽  
Pyranose Ring ◽  
Structural Requirements ◽  
Specificity Pattern

Teased renal tubules of the winter flounder (Pseudopleuronectes americanus) were employed to investigate the structural requirements for two pathways of D-glucose transport which take place preponderantly across the basal (antiluminal) face of renal cells. 1) An inhibition analysis of the equilibrating, Na-independent and phlorizin-sensitive transport of the nonmetabolizable methyl-alpha-D-glucoside (0.1 and 0.5 mM), with 20 glucose analogs (5 mM), was employed to establish the structural requirements for the substrate-carrier interaction: a (pyranose) ring, oxygen, or F at C1, C2-OH, C3-OH, and C4-OH (all axial, 1C model). Some interaction may also occur at C6-OH. D-Glucose shares this transport system. Hydrogen bonding between the oxygens and the carrier is suggested. 2) The phloretin- and phlorizin-sensitive, ouabain-insensitive transport of D-glucose, 2-deoxy-D-glucose, and D-mannose is associated with considerable phosphorylation. The three sugars mutually compete for a shared transport site. The specificity pattern characterizing the transport system defines the following structural requirements: a (pyranose) ring, a free C1-OH, C3-OH, and C4-OH (both axial) and possibly C6-OH. Hydrogen bonding between the carrier and the oxygens at C3, C4, and C6, and covalent bonding at C1 is suggested.

The Use of the Human Erythrocyte as a Model for Studying the Action of Diuretics on Sodium and Chloride Transport

1978 ◽  
Vol 54 (6) ◽  
pp. 679-683 ◽  
Author(s):  
B. A. Brooks ◽  
A. F. Lant
Keyword(s):  
Transport System ◽  
Human Erythrocyte ◽  
Chloride Transport ◽  
Transport Systems ◽  
Na Transport ◽  
Cl Transport ◽  
Henle's Loop ◽  
Low Concentrations ◽  
Biochemical Action ◽  
Chemical Analogue

1. The Na+ and Cl− transport systems of human erythrocytes have been compared for their sensitivities to diuretics known to act in the ascending limb of Henle's loop. In addition, chemical analogues of ‘loop’ compounds and also diuretics which act in other areas of the nephron have been examined. 2. The Na+ transport system lacks specificity with respect to inhibition by ‘loop’ diuretics and also a related chemical analogue studied at equivalent concentrations. 3. The Cl− transport system is inhibited, at low concentrations, by diuretics known to act in the ascending limb of Henle's loop. 4. Erythrocyte Cl− transport offers a useful model with which to study the biochemical action of diuretics.

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