GSBS Student Publications

Title

Properties of the human erythrocyte glucose transport protein are determined by cellular context

GSBS Program

Biochemistry & Molecular Pharmacology

UMMS Affiliation

Graduate School of Biomedical Sciences; Department of Biochemistry and Molecular Pharmacology

Date

4-13-2005

Document Type

Article

Medical Subject Headings

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; Transfection

Disciplines

Life Sciences | Medicine and Health Sciences

Abstract

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.

Rights and Permissions

Citation: Biochemistry. 2005 Apr 19;44(15):5606-16. Link to article on publisher's site

DOI of Published Version

10.1021/bi0477541

Related Resources

Link to article in PubMed

Journal Title

Biochemistry

PubMed ID

15823019