GSBS Dissertations and Theses
Publication Date
1991-12-01
Document Type
Doctoral Dissertation
Academic Program
Biochemistry and Molecular Pharmacology
Department
Biochemistry and Molecular Pharmacology
First Thesis Advisor
Dr. Anthony Carruthers
Keywords
Glucose Transporter Type 1
Abstract
The relationship between human erythrocyte glucose transporter (GLUT1) oligomeric structure and function was studied. GLUT1 was purified from human erythrocytes in the absence (GLUT1-DTT) or the presence (GLUT1+DTT) of dithiothreitol. Chemical cross-linking studies of lipid bilayer-resident purified GLUT1 and hydrodynamic studies of cholate-solubilized GLUT1 support the view that GLUT1-DTT is a homotetramer and GLUT1+DTT is a homodimer. Parallel studies on human erythrocyte, and studies employing conformation-specific antibodies (anti-GLUT1-DTT antibodies, ∂-IgGs), indicate that erythrocyte-resident GLUT1 resembles GLUT1-DTT (a homotetramer). While the D-glucose binding capacities of GLUT1-DTT and GLUT1+DTT are indistinguishable, GLUT1-DTT presents at least two population of binding sites to D-glucose whereas GLUT1+DTT presents only one population of sugar binding sites. The cytochalasin B (CCB) binding capacity of GLUT1-DTT (0.4 sites/monomer) is one half of that of GLUT1+DTT. GLUT1-DTT and GLUT1+DTT contain 2 and 6 free sulfhydryls per monomer respectively. The subunits (monomers) of tetrameric and dimeric GLUT1 are not linked by disulfide bridges. Erythrocyte resident GLUT1 presents at least two binding sites to D-glucose and binds CCB with a molar stoichiometry of 0.55 sites per GLUT1 monomer. Following treatment with high pH plus dithiothreitol, the sugar binding capacity of erythrocyte membrane resident transporter is unaltered but the transporter now presents only one population of binding sites to D-glucose, binds CCB with molar stoichiometry of 1.3 sites per GLUT1 monomer and displays significantly reduced affinity for ∂-IgGs. These findings demonstrate that erythrocyte resident glucose transporter is GLUT1-DTT (a GLUT1 tetramer) and that GLUT1 oligomeric structure determines GLUT1 functional properties. A model which rationalizes these findings is proposed.
Repository Citation
Hebert, DN. Glucose Transporter Oligomeric Structure Determines the Mechanism of Glucose Transport: A Dissertation. (1991). University of Massachusetts Medical School. GSBS Dissertations and Theses. Paper 217. DOI: 10.13028/1y08-kr58. https://escholarship.umassmed.edu/gsbs_diss/217
DOI
10.13028/1y08-kr58
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