Properties of the human erythrocyte glucose transport protein are determined by cellular context
UMass Chan Affiliations
Department of Biochemistry and Molecular PharmacologyGraduate School of Biomedical Sciences
Document Type
Journal ArticlePublication Date
2005-04-13Keywords
Amino Acid Substitution; Biological Transport, Active; Cell Compartmentation; Erythrocytes; Gene Deletion; Genes, Fungal; Glucose; Glucose Transporter Type 1; Humans; Kinetics; Models, Biological; Monosaccharide Transport Proteins; Mutagenesis, Site-Directed; Recombinant Proteins; Saccharomyces cerevisiae; TransfectionLife Sciences
Medicine and Health Sciences
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Show full item recordAbstract
Human erythrocyte hexose transfer is mediated by the glucose transport protein GLUT1 and is characterized by a complexity that is unexplained by available hypotheses for carrier-mediated sugar transport [Cloherty, E. K., Heard, K. S., and Carruthers, A. (1996) Biochemistry 35, 10411-10421]. The study presented here examines the possibility that the operational properties of GLUT1 are determined by host cell environment. A glucose transport-null strain of Saccharomyces cerevisiae (RE700A) was transfected with the p426 GPD yeast expression vector containing DNA encoding the wild-type human glucose transport protein (GLUT1), mutant GLUT1 (GLUT1(338)(-)(A3)), or carboxy-terminal hemagglutinin-polyHis-tagged GLUT1 (GLUT1-HA-H6). GLUT1 and GLUT1-HA-H6 are expressed at the yeast cell membrane and restore 2-deoxy-d-glucose, 3-O-methylglucose, and d-glucose transport capacity to RE700A. GLUT1-HA-H6 confers GLUT1-specific sugar transport characteristics to transfected RE700A, including inhibition by cytochalasin B and high-affinity transport of the nonmetabolized sugar 3-O-methylglucose. GLUT1(338)(-)(A3), a catalytically inactive GLUT1 mutant, is expressed but fails to restore RE700A sugar uptake capacity or growth on glucose. In contrast to transport in human red cells, K(m(app)) for 2-deoxy-d-glucose uptake equals K(i(app)) for 2-deoxy-d-glucose inhibition of 3-O-methylglucose uptake. Unlike transport in human red cells or transport in human embryonic kidney cells transfected with GLUT1-HA-H6, unidirectional sugar uptake in RE700A-GLUT1-HA-H6 is not inhibited by reductant and is not stimulated by intracellular sugar. Net uptake of subsaturating 3-O-methylglucose by RE700A-GLUT1-HA-H6 is a simple, first-order process. These findings support the hypothesis that red cell sugar transport complexity is host cell-specific.Source
Biochemistry. 2005 Apr 19;44(15):5606-16. Link to article on publisher's siteDOI
10.1021/bi0477541Permanent Link to this Item
http://hdl.handle.net/20.500.14038/33267PubMed ID
15823019Related Resources
Link to article in PubMedae974a485f413a2113503eed53cd6c53
10.1021/bi0477541