GSBS Student Publications

Title

Stop-flow analysis of cooperative interactions between GLUT1 sugar import and export sites

GSBS Program

Biochemistry & Molecular Pharmacology

UMMS Affiliation

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

Date

6-3-1999

Document Type

Article

Medical Subject Headings

3-O-Methylglucose; Binding Sites; Biological Transport; Blood Glucose; Cytochalasin B; Erythrocytes; Glucose Transporter Type 1; Humans; Kinetics; Ligands; Liposomes; Maltose; Monosaccharide Transport Proteins; Proteolipids; Spectrometry, Fluorescence

Disciplines

Life Sciences | Medicine and Health Sciences

Abstract

The human erythrocyte sugar transporter is thought to function either as a simple carrier (sugar import and sugar export sites are presented sequentially) or as a fixed-site carrier (sugar import and sugar export sites are presented simultaneously). The present study examines each hypothesis by analysis of the rapid kinetics of reversible cytochalasin B binding to the sugar export site in the presence and absence of sugars that bind to the sugar import site. Cytochalasin B binding to the purified, human erythrocyte glucose transport protein (GLUT1) induces quenching of GLUT1 intrinsic tryptophan fluorescence. The time-course of GLUT1 fluorescence quenching reflects a second-order process characterized by simple exponential kinetics. The pseudo-first-order rate constant describing fluorescence decay (kobs) increases linearly with [cytochalasin B] while the extent of fluorescence quenching increases in a saturable manner with [cytochalasin B]. Rate constants for cytochalasin B binding to GLUT1 (k1) and dissociation from the GLUT1.cytochalasin B complex (k-1) are obtained from the relationship: kobs = k-1 + k1[cytochalasin B]. Low concentrations of maltose, D-glucose, 3-O-methylglucose, and other GLUT1 import-site reactive sugars increase k-1(app) and reduce k1(app) for cytochalasin B interaction with GLUT1. Higher sugar concentrations decrease k1(app) further. The simple carrier mechanism predicts that k1(app) alone is modulated by import- and export-site reactive sugars and is thus incompatible with these findings. These results are consistent with a fixed-site carrier mechanism in which GLUT1 simultaneously presents cooperative sugar import and export sites.

Rights and Permissions

Citation: Biochemistry. 1999 May 18;38(20):6640-50. Link to article on publisher's site

DOI of Published Version

10.1021/bi990130o

Related Resources

Link to article in PubMed

Journal Title

Biochemistry

PubMed ID

10350483