Thyroxine-Binding Globulin: Characterization of the Binding Site with a Fluorescent Dye as a Probe

Abstract

The fluorescent dye 1,8-anilinonaphthalenesulfonate competed with thyroxine for binding to thyroxine-binding globulin. Fluorescence analysis indicated that the dye bound to the globulin in a molar ratio of 1:1 and with an association constant (at 23°C) of 4.19 x10⁶M^-1, and that thyroxine bound to the globulin in a molar ratio of 1:1 and with an association constant (at 23°C) of 2.35x10¹⁰M^-1. Displacement of globulin-bound dye by thyroxine was shown by fluorescence quenching, and displacement of globulin-bound thyroxine by dye was demonstrated by ultrafiltration.