Chromatographic studies of interactions between solutes, lipid bilayers and the glucose transporter Glut1
1997 (English)Doctoral thesis, comprehensive summary (Other academic)
It is advantageous to analyze solute interactions with biological membranes by chromatography on stationary phases that mimic the membrane structures. In the present work, liposomes, proteoliposomes, human red cells and red cell membrane vesicles were immobilized for analyses ofinteractions between solutes (peptides, drugs, inhibitors and glucose), lipid bilayers and theglucose transporter Glut1.
For microscale analysis of solute-membrane interactions, liposomes were immobilized incontinuous beds for capillary chromatography and included in the running buffer as pseudostationary phase in capillary electrophoresis. These techniques were convenient and rapid. Inchromatographic drug partitioning studies, the logarithm of the specific capacity factors determined on liposomes in capillary continuous beds showed a linear correlation with the logarithmof apparent drug permeabilities through Caco-2 epithelial cell monolayers.
Peptide and drug interactions with lipid bilayers were analyzed by chromatography on liposomes immobilized in gel beads. The retardations of peptides corresponding to segments ofGlut1 were related to the transfer free energy distribution within the peptides and to the hydrophobicities of the peptides. The effects of pH. temperature, ionic strength, flow rate and phospholipid composition on the retardation of drugs on liposomes or membrane vesicles were studied.
For the first time, human red cells were immobilized in gel particles for chromatographicactivity analyses. D-Glucose showed a larger retardation than L-glucose upon transport retentionchromatography and Glut1 in the cells showed high affinities for D-glucose and forskolin, asdetermined by frontal affinity analyses.
For exceptionally stable immobilization, biotin-avidin binding was introduced. Glut1 inproteoliposomes immobilized by this method showed solute affinities similar to those of sterically immobilized proteoliposomes.
Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis , 1997. , 32 p.
Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1104-232X ; 259
Biochemistry and Molecular Biology
Research subject Biochemistry
IdentifiersURN: urn:nbn:se:uu:diva-831ISBN: 91-554-3923-3OAI: oai:DiVA.org:uu-831DiVA: diva2:171017
1997-03-21, Lecture hall, B42, Biomedical Center, Uppsala University, Uppsala, 10:15