The binding of L-triiodothyronine (T3), L-thyroxine (T4), 3’-isopropyl-3, 5-diiodothyronine, 3, 5, 3’-triiodothyroacetic acid, 3, 3’, 5’-triiodothyronine, and sulfobromophthailine (BSP) by rat liver and T3, T4 and BSP by kidney cytosol was analyzed in displacement experiments using tracer 125I-T3, graded doses of nonradioactive ligand, and dextrancoated charcoal to separate bound and free ligands. Plots of the ratio of 125I-T3 bound to cytosol (bound) to the 125I-T3 in charcoal (free) against the concentration of cytosol (bound) 125I-T3 revealed two classes of T3 binding sites: (A) a set of nonspecific sites with low affinity and apparently unlimited capacity, and (B) a set of limited capacity sites with higher affinity. In liver cytosol the equilibrium constant of the latter set was 4.3 × 107M-1 and the binding capacity was 5.3 × 107M. The corresponding values in kidney cytosol were 2.5 × 107M-1 and 2.9 × 10-6M. Thyroxine, all of the T3 analogues used, and BSP appeared to bind to the same sites with a lesser affinity. The observed T3 binding characteristics of cytosol protein appeared to differ markedly from those recently described for the nucleus. First, the apparent association constant of the cytosol T3 binding sites was less than l/200th that of the nucleus. Secondly, the nuclear T3 binding sites appear to be at least 70% saturated at endogenous T3 concentrations in euthyroid animals, whereas less than 1% of cytosol sites are occupied by T3. Finally, the relative strength of binding of T3 analogues to cytosol differs markedly from the corresponding pattern at the nuclear site. These differences indicate that nuclear binding of T3 cannot represent simple translocation of cytosol T3 receptor sites to the nucleus. Either different sites are involved or the binding characteristics of the cytosol protein T3 complex are markedly altered once the putative T3 receptor enters the nucleus.
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