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

Barnett, J. E. G.; Holman, Geoffrey D.; Munday, K. A.

doi:10.1042/bj1350539

Cited by 48 publications

(24 citation statements)

References 16 publications

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“…The observations argue for some rearrangement of the substrate binding region during carrier reorientation. Dissimilar specificity at inner and outer transport sites is seen in other systems, for example the glucose and choline carriers of red cells, where even close analogs of the substrate may be bound exclusively on one side of the membrane (Barnett, Holman & Munday, 1973;Devds & Krupka, 1984).…”

Section: Gomentioning

confidence: 97%

Role of substrate binding forces in exchange-only transport systems: II. Implications for the mechanism of the anion exchanger of red cells

1989

J. Membrain Biol.

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The transition-state theory of exchange-only membrane transport is applied to experimental results in the literature on the anion exchanger of red cells. Two central features of the system are in accord with the theory: (i) forming the transition state in translocation involves a carrier conformational change; (ii) substrate specificity is expressed in transport rates rather than affinities. The expression of specificity is consistent with other evidence for a conformational intermediate (not the transition state) formed in the translocation of all substrates. The theory, in conjunction with concepts derived from the chemistry of macrocyclic ion inclusion complexes, prescribes certain essential properties in the transport site. Separate subsites are required for the preferred substrates, Cl- and HCO3-, to account for tight binding in the transition state (Kdiss congruent to 1 microM). Further, the following mechanism is suggested. A substrate anion initially forms a loose surface complex at one subsite, but in the transition state the subsites converge to form an inclusion complex in which the binding forces are greatly increased through a chelation effect. The conformational change at the substrate site, which is driven by the mounting forces of binding, sets in train a wider conformational change that converts the carrier from an immobile to a mobile form. Though simple, this composite-site mechanism explains many unusual features of the system. It accounts for substrate inhibition, partially noncompetitive inhibition of one substrate by another, and "tunneling," which is net transport under conditions where exchange should prevail, according to other models. All three types of behavior result from the formation of a ternary complex in which substrate anions are bound at both subsites. The mechanism also accounts for the enormous range of substrate structures accepted by the system, for the complex inhibition by the organic sulfate NAP-taurine, and for the involvement of several cationic side chains and two different protein domains in the transport site.

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Section: Gomentioning

confidence: 97%

Role of substrate binding forces in exchange-only transport systems: II. Implications for the mechanism of the anion exchanger of red cells

1989

J. Membrain Biol.

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“…The observation that derivatives of glucose substituted at C-4 and C-6 were preferentially bound outside, while a derivative substituted at C-1 was preferentially bound inside, was explained by Barnett et al [4] on the basis of a single substrate site, part of which binds the substrate on one surface, and part on the other: at the outer site C-1 participates in binding, and C-4 and C-6 are in contact with the solvent, while at the inner site the reverse is true. Doubt was thrown on this hypothesis by the later report of Baker et al [2], that two other analogs substiuted at the C-1 end of the glucose molecule are more firmly bound outside than inside.…”

Section: Discussionmentioning

confidence: 86%

“…Binding asymmetry has also been observed in the glucose carrier of human erythrocytes [2][3][4]. The observation that derivatives of glucose substituted at C-4 and C-6 were preferentially bound outside, while a derivative substituted at C-1 was preferentially bound inside, was explained by Barnett et al [4] on the basis of a single substrate site, part of which binds the substrate on one surface, and part on the other: at the outer site C-1 participates in binding, and C-4 and C-6 are in contact with the solvent, while at the inner site the reverse is true.…”

Section: Discussionmentioning

confidence: 92%

The comparative specificity of the inner and outer substrate transfer sites in the choline carrier of human erythrocytes

Devés

Krupka

1984

J. Membrain Biol.

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The substrate specificities on the inner and outer surfaces of the cell membrane have been compared by determining the relative affinities, inside and outside, of a series of choline analogs. The results of two different methods were in agreement: (1) the carrier distribution was determined in the presence of a saturating concentration of an equilibrated analog, using N-ethylmaleimide as a probe for the inward-facing carrier; (2) the degree of competition was measured between an equilibrated analog and choline in the external solution. The carrier sites are found to have markedly different specificities: the outer site is more closely complementary to the structure of choline than is the inner, and even a slight enlargement of either the trimethylammonium or hydroxyethyl group gives rise to preferential binding inside. It is also found that a nonpolar binding region, which is adjacent to the outer site, is absent from the inner site. As the transport mechanism involves the exposure of only one site at a time, first on one surface and then the other, it follows that an extensive reorganization of the structure of the substrate site may occur during the carrier-reorientation step, or alternatively that two distinct sites may be present, only one of which is exposed at a time.

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“…Previous studies have established that bulky groups are tolerated around the 4-OH position when the hexose occupies the exofacial binding sites of the transporters. 15) The bis-hexose compounds we have developed are linked by a bridge through their C-4 positions. The remaining hexose hydroxyls are available for interaction with the GLUT4 protein.…”

Section: Resultsmentioning

confidence: 99%

Improvement in the Properties of 3-Phenyl-3-trifluoromethyldiazirine Based Photoreactive Bis-Glucose Probes for GLUT4 Following Substitution on the Phenyl Ring.

Hashimoto

Yang

Hatanaka

et al. 2002

CHEMICAL & PHARMACEUTICAL BULLETIN

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Recent human genome sequencing has led to the realisation that the family of facilitative sugar transporters (GLUTs: glucose transporters) in mammals is more extensive than the group of five members (GLUTs 1-5) described a decade ago. Now the GLUTs are known to be a more complex family of thirteen isomeric proteins that is divisible into three subgroups based on sequence similarities. 1) Class 1 consists of the glucose transporters (GLUTs 1-4) which preferentially transport D-glucose. 2) Of these transporters GLUT4 is particularly important both in physiological and pathophysiological processes as it is present only in insulin-responsive tissues such as adipose, heart and skeletal muscle. Following insulin signaling, there is an increase in GLUT4 exocytosis from intracellular storage vesicles and incorporation of the protein into the surface membrane of the insulin target cells. 3) Loss of responsiveness of this protein to insulin is one of the contributing factors to the pathogenesis of Type 2 diabetes. 4) To effectively examine the mechanisms involved in exposure of GLUT4 at the cell surface of adipocytes, high affinity analogues that can tag the transporter are required. Synthesizing suitable analogues is a difficult task since sugars have relatively low affinity for the transporter molecules. D-Glucose for example, when interacting with GLUT4, does so with an affinity constant (K m ) of around 8 mM. Physiologically the high K m is important as if the affinity for D-glucose were very high then this substrate would bind very strongly to the transporter rather than being transported through it. In addition, blood glucose levels are generally 5 mM (or slightly higher following a carbohydrate rich meal) and therefore a high affinity (low K m ) system would become too easily saturated and unable to respond to the fluctuations in circulating blood glucose levels. The requirements for a GLUT4 tag are opposite to these and high affinity interaction is required in a photolabel as the ligand/protein complex has to be occupied at a high enough level to be efficiently converted to a covalent complex following UV irradiation. 5) We have developed a series of photoaffinity probes based on bis-hexose structures. 6) These compounds contain hexoses linked via their 4-OH positions to a propyl-2-amine spacer. Because the bis-hexose structure renders the compounds large and hydrophilic the derivatives are impermeant and just probe those glucose transporters (GLUT4) that appear at the cell-surface in response to insulin. We have already synthesized photoreactive bis-mannose derivatives containing arylazide, 7) benzophenone 8) and 3-phenyl-3-trifluoromethyldiazirine 9) substitutions for use in glucose transporter cell surface labeling. Recently, we have synthesized photoreactive bis-glucose derivatives containing the 3-phenyl-3-trifluoromethyldiazirine group. 10) These can easily be synthesised preparative scale and have slightly higher affinity for GLUT4 than the equivalent bis-mannose compounds.Arylazide ligands were found to b...

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

Cited by 48 publications

References 16 publications

Role of substrate binding forces in exchange-only transport systems: II. Implications for the mechanism of the anion exchanger of red cells

Role of substrate binding forces in exchange-only transport systems: II. Implications for the mechanism of the anion exchanger of red cells

The comparative specificity of the inner and outer substrate transfer sites in the choline carrier of human erythrocytes

Improvement in the Properties of 3-Phenyl-3-trifluoromethyldiazirine Based Photoreactive Bis-Glucose Probes for GLUT4 Following Substitution on the Phenyl Ring.

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