Demonstration of specific binding sites for 3H-RRR-alpha-tocopherol on human erythrocytes.
Previous work from our laboratory demonstrated specific binding sites for 3H-RRR-alpha-tocopherol (3H-d alpha T) in membranes of rat adrenal cells. As tocopherol deficiency is associated with increased susceptibility of red blood cells to hemolysis, we investigated tocopherol binding sites in human RBCs. Erythrocytes were found to have specific binding sites for 3H-d alpha T that exhibited saturability and time and cell-concentration dependence as well as reversibility of binding. Kinetic studies of binding demonstrated two binding sites--one with high affinity (Ka of 2.6 x 10(7) M-1), low capacity (7,600 sites per cell) and the other with low affinity (1.2 x 10(6) M-1), high capacity (150,000 sites per cell). In order to localize the binding sites further, RBCs were fractionated and greater than 90% of the tocopherol binding was located in the membranes. Similar to the findings in intact RBCs, the membranes exhibited two binding sites with a respective Ka of 3.3 x 10(7) M-1 and 1.5 x 10(6) M-1. Specificity data for binding demonstrated 10% binding for RRR-gamma-tocopherol, but not other tocopherol analog exhibited competition for 3H-d alpha T binding sites. Instability data suggested a protein nature for these binding sites. Preliminary studies on Triton X-100 solubilized fractions resolved the binding sites to a major component with an Mr of 65,000 and a minor component with an Mr of 125,000. We conclude that human erythrocyte membranes contain specific binding sites for RRR-alpha-tocopherol. These sites may be of physiologic significance in the function of tocopherol on the red blood cell membrane.
D004910:Erythrocyte Membrane / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D006461:Hemolysis; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism*; D011973:Receptors, LDL; D014316:Tritium; D014810:Vitamin E / Q000378:metabolism*
D011956:Receptors, Cell Surface; D011973:Receptors, LDL; C027656:alpha-tocopherol receptor; D014316:Tritium; D014810:Vitamin E