The Binding Stoichiometry of Human Plasma Retinol-binding Protein to Prealbumin

The stoichiometry of binding between all-trans-retinol-retinol-binding protein (RBP) and prealbumin was investigated by means of gel filtration chromatography, electrophoresis, absorption, and circular dichroism. Using protein concentrations of up to 20 mg per ml, it was found that under a variety of experimental conditions 1 prealbumin molecule binds 1 to 1.35 molecules of retinol-RBP at pH 7.0 in an 0.033 m sodium phosphate buffer containing 0.1 m NaCl. These results were further supported by hybridization experiments using both all-trans-retinol-RBP and all-trans-3-dehydroretinol-RBP and prealbumin. It was found that the free chromophore-RBP molecules were freely exchangeable with the chromophore-RBP molecules which were bound to prealbumin, but the sum total of chromophore-RBP bound to prealbumin was 1:1 on a molar basis.