Negative cooperativity in the binding of thyroxine to human serum prealbumin

Abstract

The binding of thyroxine (T4) and 8-anilino-1-naphthalenesulfonic acid (ANS) to human serum prealbumin was measured by equilibrium dialysis at pH 7.4 in 0.05 M phosphate-0.10 M NaCl at 25 degrees. The data were analyzed for the binding constants based on equations for (1) two independent sites and (2) two identical sites with negative interaction. Evaluation by the independent site model gave the following association constants: for T4 binding, KT1 = 1.0 x 10-8 M-1, KT2 = 9.5 x 10-5 M-1; for ANS binding, KA1 = 9.5 x 10-5 M-1, KA2 = 2.1 x 10-5 M-1. The interactive model gave constants kT = 5.5 x 10-7 M-1 and kA = 5.5 x 10-5 M-1. Interaction factors, alpha, defined such that -RT in alpha is the energy of interaction, were: alpha T = 0.041 AND ALPHA A = 0.62 for T4 and ANS, respectively. The "best fit" values for the number of sites were 2.0 and 1.6 for T4 and ANS, respectively. The binding of T4 to human prealbumin was competitive with ANS, and the binding constants evaluated from competition experiments were in agreement with those found for each ligand when studied separately. On the basis of analysis of X-ray data of human prealbumin (Blake et al.) there appear to be two identical T4 sites. It is therefore evident that the binding of T4 represents a case of negative cooperativity which is presumably due to interaction between ligands.

The Crystallographic Structure of Phytohemagglutinin-L

“…) decreases the affinity of the other site (10 6 M Ϫ1 ) (Ferguson et al, 1975). Again as in PHA-L, these sites show internal 2-fold symmetry.…”

The evolutionary and integrative roles of transthyretin in thyroid hormone homeostasis

“…The tetramer contains a central channel with the two binding sites for thyroid hormones (Blake et al 1978). There is negative cooperativity between the two sites (Ferguson et al 1975, Neumann et al 2001. The best conserved regions of the polypeptide chain are the binding site and the amino acids lining the channel , Prapunpoj et al 2000a.…”

Fluorescence study of the thyroxine‐dependent conformational changes in human serum transthyretin

“…Two mol of T4 are bound per mol of protein with binding constants two orders of magnitude different for each tool. This is interpreted in terms of a negative cooperativity between the sites [6,7]. The study described here was undertaken with the intention of using TTR Trp-41 and Trp-79 fluorescence which can give structural information, to detect conformational changes associated to its interaction with T4.…”