The heterozygous substitution of threonine for alanine at amino acid 109 of human transthyretin (TTR) increases its affinity for T4. We compared the affinity of recombinant wild-type (WT) and Thr109-TTRs for various iodothyronines in an attempt to elucidate how this mutation alters the T4- binding site. Homozygous WT and Thr109-TTRs were expressed recombinantly in Escherichia coli, and heterozygous Thr109-TTR was purified from plasma. The affinities of the iodothyronines for TTR were determined by measuring [125I]T4 bound by TTR in the presence of increasing concentrations of unlabeled iodothyronines. Homozygous Thr109-TTR bound T4 with an affinity slightly, but not significantly, greater than that of heterozygous Thr109- TTR. The affinity of Thr109-TTR for all iodothyronines was higher than that of WT TTR. However, the Thr109 mutation increased TTR's affinity for T4, Triac (triiodothyroacetic acid), and T3 to a greater extent than it did for Tetrac (tetraiodothyroacetic acid), EMD21388 (3',5'-dibromo-4',6'- dihydroxy-3-methylflavone), and dextro-T4. These data demonstrate that a subtle change in the structure of the T4-binding channel in TTR differentially alters the affinity of binding of various iodothyronines and suggests that site-directed mutagenesis of residues within the binding channel might clarify the relative importance of specific domains of this binding channel.
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