Linked Life Data

1517202

Source:http://linkedlifedata.com/resource/pubmed/id/1517202

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
25
pubmed:dateCreated
1992-10-7
pubmed:abstractText
The structure-function relationship of the HepG2/erythrocyte-type glucose transporter (GLUT1) has been studied by in vitro site-directed mutagenesis. Chinese hamster ovary clones in which glucose transporters were transfected were shown by Western blotting with a GLUT1 anti-COOH-terminal peptide antibody to have expression levels of Gln282----Leu, Asn288----Ile, and Asn317----Ile mutations that were comparable with the wild type. All three mutant GLUT1 clones had high 2-deoxy-D-glucose transport activity compared with a nontransfected clone, suggesting that these residues are not absolutely required for the transport function. We have examined the possibility that the inner and outer portions of the transport pathway are structurally separate by measuring the interaction of the mutant transporters with the inside site-specific ligand cytochalasin B and the outside site-specific ligand 2-N-4-(1-azi-2,2,2-trifluoroethyl)benzoyl-1,3-bis(D-mannos-4 -yloxy)-2- propylamine (ATB-BMPA). All three mutant GLUT1 clones showed high levels of cytochalasin B labeling, and the N288I and N317I mutants showed high levels of ATB-BMPA labeling. In contrast to the transport and cytochalasin B labeling results, the transmembrane helix 7 Gln282----Leu mutant was labeled by ATB-BMPA to a level that was only 5% of the level observed in the wild type. We have confirmed that this mutant was defective in the outer site by comparing the inhibition of wild-type and mutant 2-deoxy-D-glucose transport by the outside site-specific ligand 4,6-O-ethylidene-D-glucose. 4,6-O-Ethylidene-D-glucose inhibited wild-type transport with a Ki of approximately 12 mM, but this was increased to greater than 120 mM in the Gln282----Leu mutant. Thus, of the 3 residues mutated in this study, only glutamine 282 substitution causes a major perturbation in function, and this is a specific and striking reduction in the affinity for the outside site-specific ligands ATB-BMPA and 4,6-O-ethylidene-D-glucose.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Sep
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
5
pubmed:volume
267
pubmed:geneSymbol
GLUT1
pubmed:owner
NLM
pubmed:authorsComplete
N
pubmed:pagination
17502-7
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed-meshheading:1517202-Amino Acid Sequence, pubmed-meshheading:1517202-Animals, pubmed-meshheading:1517202-Antibodies, pubmed-meshheading:1517202-Base Sequence, pubmed-meshheading:1517202-Binding Sites, pubmed-meshheading:1517202-Biological Transport, pubmed-meshheading:1517202-CHO Cells, pubmed-meshheading:1517202-Cloning, Molecular, pubmed-meshheading:1517202-Cricetinae, pubmed-meshheading:1517202-Genetic Vectors, pubmed-meshheading:1517202-Glucose, pubmed-meshheading:1517202-Humans, pubmed-meshheading:1517202-Kinetics, pubmed-meshheading:1517202-Molecular Sequence Data, pubmed-meshheading:1517202-Monosaccharide Transport Proteins, pubmed-meshheading:1517202-Mutagenesis, Site-Directed, pubmed-meshheading:1517202-Oligodeoxyribonucleotides, pubmed-meshheading:1517202-Oligopeptides, pubmed-meshheading:1517202-Protein Conformation, pubmed-meshheading:1517202-Recombinant Proteins, pubmed-meshheading:1517202-Restriction Mapping, pubmed-meshheading:1517202-Transfection
pubmed:year
1992
pubmed:articleTitle
Site-directed mutagenesis of GLUT1 in helix 7 residue 282 results in perturbation of exofacial ligand binding.
pubmed:affiliation
Second Department of Internal Medicine, Kobe University School of Medicine, Japan.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't