High affinity phlorizin binding to the LLC-PK1 cells exhibits a sodium:phlorizin stoichiometry of 2:1.
The phlorizin binding properties of luminal membrane vesicles isolated from the LLC-PK1 cells, a continuous epithelial cell line derived from pig kidney, are studied. Scatchard analysis of this binding indicates the existence of a single high affinity sodium-dependent site with KD = 0.4 microM at 266 mM sodium. The specificity properties of this site indicate that it represents the binding of phlorizin to the hexose binding site of the sodium-dependent D-glucose transporter previously identified in this cell line. Both phlorizin equilibrium binding and the rate of phlorizin binding were found to be sigmoidal functions of sodium concentration. A Hill analysis of these data was consistent with a sodium:phlorizin stoichiometry of 2:1 in good agreement with the sodium:glucose stoichiometry already established in these cells. Phlorizin dissociation was also found to be sodium-dependent. On the basis of the phlorizin binding data presented here, a number of models of the binding of phlorizin and sodium to the transporter can be excluded. An analysis of a random binding model consistent with the data is presented. The significance of the LLC-PK1 sodium-dependent D-glucose transporter as a model system for related renal and intestinal transporters is discussed.
Moran
A
A
Department of Physiology, Armed Forces Radiobiological Research Institute, Bethesda, Maryland 20892.
Davis
L J
LJ
Turner
R J
RJ
D000818:Animals; D002460:Cell Line; D007700:Kinetics; D008871:Microvilli / Q000378:metabolism*; D008954:Models, Biological; D010695:Phlorhizin / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*
D012964:Sodium; D010695:Phlorhizin