Relevance of the Low and High Affinity Thyrotropin-Binding Sites of Human Thyroid Membranes to the Stimulation of Adenylate Cyclase

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Abstract
TSH is known to interact on thyroid membranes with 2 classes of binding sites that differ in affinity and capacity. To assess the relevance of the class of TSH-binding sites characterized by low affinity and high capacity to the stimulation of adenylate cyclase, the interactions of desialylated hCG [human chorionic gonadotropin] (as-hCG) and its .beta.-subunit (as-hCG.beta.) with human thyroid membranes were studied. In low ionic strength buffer, pH 7.8, where both classes of sites are operant, as-hCG fully inhibited and as-hCG.beta. partially inhibited [125I] bovine (b) TSH binding. Scatchard analysis of the [125I]bTSH binding inhibition curve in the presence of 1.0 .times. 10-5 M as-hCG.beta. clearly indicated that as-hCG.beta. interacted only with the low affinity class of binding sites, leaving the high affinity class unaffected. In the presence of 140 mM NaCl, [125I]bTSH interacted predominantly with the high affinity class of binding sites; as-hCG fully inhibited [125I]bTSH binding to this class of sites; as-hCG.beta. displayed essentially no interaction. Scatchard analysis of [125I]as-hCG.beta. binding to human thyroid membranes in low ionic strength buffer revealed a single apparent class of sites with low affinity (Kd = .apprx. 1.0 .times. 10-6 M) and high capacity (Q = .apprx. 300 pmol/mg membrane protein). The bTSH preparation (Thytropar) showed a 10-fold greater binding inhibition potency at these sites than either the as-hCG or the as-hCG.beta. preparation, in keeping with the inference that as-hCG.beta. interacts with the low affinity class of TSH-binding sites. At a concentration > 3 times that necessary to inhibit TSH binding to the low affinity class of sites, the as-hCG.beta. molecule neither stimulated adenylate cyclase nor inhibited the ability of TSH to do so. In contrast, the as-hCG molecule, which interacts with both classes of TSH-binding sites, fully inhibited TSH stimulation of adenylate cyclase. The low affinity class of TSH-binding sites is not the class of sites through which TSH stimulates adenylate cyclase, and this role is best ascribed to the high affinity class of TSH-binding sites.