Thyroxine-Binding Globulin: Characterization of the Binding Site with a Fluorescent Dye as a Probe
Allan M. Green 1,
James S. Marshall 2,
Jack Pensky 2, and
John B. Stanbury 3
1 Department of Medicine, Case Western Reserve University, Cleveland, Ohio 44106, and Department of Nutrition, Massachusetts Institute of Technology, Cambridge 02139
2 Department of Medicine, Case Western University
3 Department of Nutrition, Massachusetts Institute of Technology
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.
