IN-VITRO BINDING OF 3,5-DI-IODO-L-THYRONINE TO RAT-LIVER MITOCHONDRIA

In this study we have demonstrated that specific binding sites for 3,5-di-iodo-L-thyronine (3,5-T2) can be detected in rat liver mitochondria. After incubation with the homogenate, liver mitochondria bound only a small portion of [3,5,-125I]T2. The addition of a 100-fold excess of unlabelled 3,5-T2 caused the displacement of on average 50-60% of the [3,5-125I]T2 bound. Specific binding of 3,5-T2 to rat fiver mitochondria occurred rapidly; a maximum was achieved after 5 min. Maximal binding was obtained at 37°C, while at 0°C and 20°C the values were only slightly lower. Binding was maximal at pH 7.0; mean (±S.E.M.) values for the apparent association constant and the binding capacity (calculated at pH 7.0, 0°C and after 30 min of incubation) were 0.5 ± 0.04 x 108 M-1 and 0.4 ± 0.04 pmol/mg mitochondrial protein respectively. The specificity of binding, examined in competition studies, followed the order: 3,5-T2> 3,3'-di-iodo-L-thyronine> 3',3,5-tri-iodo-L-thyronine>thyroxine. Other iodothyronines (3',5'-di-iodo-L-thyronine, 3,5-di-iodo-D-thyronine, 3,3',5'-tri-iodo-L-thyronine, 3-iodo-L-thyronine and 3,5-di-iodothyroacetic acid) showed little or no competition. This suggests that the specific 3,5-T2 binding sites could be of biological relevance with respect to the understanding of the mechanism of physibiogical action of thyroid hormones at the cellular level.