Phlorizin receptors in isolated kidney brush border membranes. Differential enzymatic modification of high-affinity receptors and unspecific binding sites.

Isolated brush border membranes from rat kidney bind 13H]phlorizin specifically. A “high affinity” binding site (Ka (the association constant) = 5.0 X lo6 1 X mole-r at 150 mu sodium, pH 7.4,37”) is enriched in the brush border membrane and is not present in kidney mitochondria or red blood cell ghosts of the rat. The concentration of the high affinity receptors in the membrane fraction is 40 pmoles per mg of protein, about 17 times higher than in a kidney homogenate. Inhibition studies with monosaccharides demonstrate that the binding curve is heterogeneous. D-Glucose, D-galactose, and Z-deoxy-D-glucose inhibited phlorizin binding at the high atl’inity site, whereas D-fructose, D-mannose, and L-arabinose interact with another “low affinity” receptor in the brush border membrane. The inhibition studies show that its affinity for phlorizin is one to two orders of magnitude lower than that of the high affinity receptor. The high affinity receptors are destroyed by treatment with trypsin and sulfhydryl reagents like p-hydroxymercuriphenylsulfonic acid and N-ethylmaleimide. Phlorizin binding is sodium-dependent; the affinity of the high affinity receptor decreases 30-fold when the sodium concentration is lowered from 150 to 10 rnM. Kidney brush border membranes contain no detectable phlorizin hydrolase, but the receptors are rapidly degraded by a temperature-dependent process that can be inhibited by ethylenediaminetetraacetate. The stereospecificity of the phlorizin receptors in the isolated luminal membrane of the rat kidney proximal tubule has remarkable similarity to the stereospecificity of the monosaccharide carriers as shown by in vivo experiments (SILVERMAN, M., AGANON, M. A.,AND CHINARD, P. F. (1970) Amer. J. Physiol. 218, 735; 743).