Thyroid hormone binding by recombinant sea bream transthyretin: the role of the N-terminal region

Transthyretin (TTR) along with thyroxine-binding globulin (TBG) and albumin (ALB) constitute Thyroid Hormone-Binding Proteins (THBP) in vertebrates. These proteins bind and transport thyroid hormones (THs), thyroxine (T4) and triiodothyronine (T3) in the blood. THBP are poorly characterized in fish and in the present study binding of THs by sea bream TTR was determined. Teleost TTR is a single polypeptide chain of 130 amino acids, but in common with mammals the functional form in the plasma is a tetramer. In contrast to mammalian TTR (127 amino acids), fish TTR has a longer N-terminal region and the latter has been proposed to influence TH binding. In the present study recombinant sea bream TTR (sbTTR, wild type), plus two recombinant N-terminal mutants were produced and purified. Binding of [I125]-T3 to the purified TTRs was confirmed by polyacrylamide gel electrophoresis under nondenaturing conditions followed by autoradiography and confirmed all bind THs as a tetramer. Ligand binding studies with labeled [I125]-T3 were performed and revealed that [I125]-T3 was displaced in a similar way from wild type and mutant TTRs by increasing concentrations of unlabeled T3. Similar Kd values were obtained for both T3 and T4 binding to wild type and mutant TTRs indicating that the N-terminal region does not seem to be important for the binding characteristics of sbTTR