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dc.contributor.authorSage, Jay M.
dc.contributor.authorCura, Anthony J.
dc.contributor.authorLloyd, Kenneth P.
dc.contributor.authorCarruthers, Anthony
dc.date2022-08-11T08:08:55.000
dc.date.accessioned2022-08-23T16:12:22Z
dc.date.available2022-08-23T16:12:22Z
dc.date.issued2015-05-15
dc.date.submitted2015-08-31
dc.identifier.citationAm J Physiol Cell Physiol. 2015 May 15;308(10):C827-34. doi: 10.1152/ajpcell.00001.2015. Epub 2015 Feb 25. <a href="http://dx.doi.org/10.1152/ajpcell.00001.2015">Link to article on publisher's site</a>
dc.identifier.issn0363-6143 (Linking)
dc.identifier.doi10.1152/ajpcell.00001.2015
dc.identifier.pmid25715702
dc.identifier.urihttp://hdl.handle.net/20.500.14038/33381
dc.description.abstractGlucose transporter 1 (GLUT1) is the primary glucose transport protein of the cardiovascular system and astroglia. A recent study proposes that caffeine uncompetitive inhibition of GLUT1 results from interactions at an exofacial GLUT1 site. Intracellular ATP is also an uncompetitive GLUT1 inhibitor and shares structural similarities with caffeine, suggesting that caffeine acts at the previously characterized endofacial GLUT1 nucleotide-binding site. We tested this by confirming that caffeine uncompetitively inhibits GLUT1-mediated 3-O-methylglucose uptake in human erythrocytes [Vmax and Km for transport are reduced fourfold; Ki(app) = 3.5 mM caffeine]. ATP and AMP antagonize caffeine inhibition of 3-O-methylglucose uptake in erythrocyte ghosts by increasing Ki(app) for caffeine inhibition of transport from 0.9 +/- 0.3 mM in the absence of intracellular nucleotides to 2.6 +/- 0.6 and 2.4 +/- 0.5 mM in the presence of 5 mM intracellular ATP or AMP, respectively. Extracellular ATP has no effect on sugar uptake or its inhibition by caffeine. Caffeine and ATP displace the fluorescent ATP derivative, trinitrophenyl-ATP, from the GLUT1 nucleotide-binding site, but d-glucose and the transport inhibitor cytochalasin B do not. Caffeine, but not ATP, inhibits cytochalasin B binding to GLUT1. Like ATP, caffeine renders the GLUT1 carboxy-terminus less accessible to peptide-directed antibodies, but cytochalasin B and d-glucose do not. These results suggest that the caffeine-binding site bridges two nonoverlapping GLUT1 endofacial sites-the regulatory, nucleotide-binding site and the cytochalasin B-binding site. Caffeine binding to GLUT1 mimics the action of ATP but not cytochalasin B on sugar transport. Molecular docking studies support this hypothesis.
dc.language.isoen_US
dc.relation<a href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Retrieve&list_uids=25715702&dopt=Abstract">Link to Article in PubMed</a>
dc.relation.urlhttp://dx.doi.org/10.1152/ajpcell.00001.2015
dc.subject3-O-Methylglucose; Adenosine Triphosphate; Binding Sites; Biological Transport; Caffeine; Cytochalasin B; Erythrocyte Membrane; Erythrocytes; Glucose; Glucose Transporter Type 1; Humans; Signal Transduction
dc.subjectBiochemistry
dc.subjectCellular and Molecular Physiology
dc.titleCaffeine inhibits glucose transport by binding at the GLUT1 nucleotide-binding site
dc.typeJournal Article
dc.source.journaltitleAmerican journal of physiology. Cell physiology
dc.source.volume308
dc.source.issue10
dc.identifier.legacycoverpagehttps://escholarship.umassmed.edu/gsbs_sp/1906
dc.identifier.contextkey7536280
html.description.abstract<p>Glucose transporter 1 (GLUT1) is the primary glucose transport protein of the cardiovascular system and astroglia. A recent study proposes that caffeine uncompetitive inhibition of GLUT1 results from interactions at an exofacial GLUT1 site. Intracellular ATP is also an uncompetitive GLUT1 inhibitor and shares structural similarities with caffeine, suggesting that caffeine acts at the previously characterized endofacial GLUT1 nucleotide-binding site. We tested this by confirming that caffeine uncompetitively inhibits GLUT1-mediated 3-O-methylglucose uptake in human erythrocytes [Vmax and Km for transport are reduced fourfold; Ki(app) = 3.5 mM caffeine]. ATP and AMP antagonize caffeine inhibition of 3-O-methylglucose uptake in erythrocyte ghosts by increasing Ki(app) for caffeine inhibition of transport from 0.9 +/- 0.3 mM in the absence of intracellular nucleotides to 2.6 +/- 0.6 and 2.4 +/- 0.5 mM in the presence of 5 mM intracellular ATP or AMP, respectively. Extracellular ATP has no effect on sugar uptake or its inhibition by caffeine. Caffeine and ATP displace the fluorescent ATP derivative, trinitrophenyl-ATP, from the GLUT1 nucleotide-binding site, but d-glucose and the transport inhibitor cytochalasin B do not. Caffeine, but not ATP, inhibits cytochalasin B binding to GLUT1. Like ATP, caffeine renders the GLUT1 carboxy-terminus less accessible to peptide-directed antibodies, but cytochalasin B and d-glucose do not. These results suggest that the caffeine-binding site bridges two nonoverlapping GLUT1 endofacial sites-the regulatory, nucleotide-binding site and the cytochalasin B-binding site. Caffeine binding to GLUT1 mimics the action of ATP but not cytochalasin B on sugar transport. Molecular docking studies support this hypothesis.</p>
dc.identifier.submissionpathgsbs_sp/1906
dc.contributor.departmentDepartment of Biochemistry and Molecular Pharmacology
dc.source.pagesC827-34
dc.contributor.studentKenneth P. Lloyd


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