Electron spin resonance investigation of the interaction of the anion and glucose transport inhibitor, p-azidobenzylphlorizin, with the human red cell membrane.
The membrane perturbations caused by the interaction of p-azidobenzylphlorizin (p-AzBPhz), a potential photoaffinity labeling agent of the anion and D-glucose transporters in the human erythrocyte, have been studied using electron spin resonance (ESR) spectrometry. Two lipid-specific spin labels have been employed; one of these agents, a hexadecyl-quarternary amine with the nitroxide reporter group covalently attached to the cationic nitrogen, (CAT-16), has been used to monitor changes in the physical state of the membrane's extracellular phospholipid/water interface. The other spin label, 5-doxylstearic acid (5-NS), is designed to examine the order and motion of the lipid bilayer near the cell surface. In separate experiments, intact human red cells labeled with these lipid-specific spin labels were exposed to small amounts of the phlorizin azide. A dose-dependent alteration in CAT-16 motion was observed, but the p-AzBPhz interaction with the membrane had no effect on the spectrum of 5-NS. The half-maximal effect of the phlorizin derivative on the CAT-16 spectrum occurred when about 2 million molecules were bound to each cell. This is also the combined amount of band 3 and band 4.5 present in the red cell membrane and represents the concentration necessary to inhibit both anion and glucose transport. Our results suggest that the first p-AzBPhz molecules binding to the red cell membrane interact with the anion and sugar transporters, and not with the bulk lipid bilayer.
Wyse
J W
JW
Center of Membrane Sciences, University of Kentucky, Lexington 40506-0055.
Blank
M E
ME
Maynard
C L
CL
Diedrich
D F
DF
Butterfield
D A
DA
D027321:Anion Transport Proteins; D001386:Azides / Q000494:pharmacology*; D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000037:antagonists & inhibitors* / Q000097:blood; D002712:Chlorides / Q000097:blood; D004578:Electron Spin Resonance Spectroscopy; D004910:Erythrocyte Membrane / Q000187:drug effects*; D006801:Humans; D066298:In Vitro Techniques; D008051:Lipid Bilayers; D008560:Membrane Fluidity; D009004:Monosaccharide Transport Proteins / Q000037:antagonists & inhibitors* / Q000097:blood; D009068:Movement; D010695:Phlorhizin / Q000031:analogs & derivatives* / Q000494:pharmacology
D027321:Anion Transport Proteins; D001386:Azides; D002352:Carrier Proteins; D002712:Chlorides; D008051:Lipid Bilayers; D009004:Monosaccharide Transport Proteins; C056185:phlorizinyl 5'-benzylazide; D010695:Phlorhizin