title: Relevance of the low and high affinity thyrotropin-binding sites of human thyroid membranes to the stimulation of adenylate cyclase.
abstract: TSH is known to interact on thyroid membranes with two 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, we studied the interactions of desialylated hCG (as-hCG) and its beta-subunit (as-hCG beta) with human thyroid membranes. 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 X 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, whereas 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 = approximately 1.0 X 10(-6) M) and high capacity (Q = approximately 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 more than 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. We conclude that the low affinity class of TSH-binding sites is not the class of sites through which TSH stimulates adenylate cyclase, and that this role is best ascribed to the high affinity class of TSH-binding sites.
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism*; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D006063:Chorionic Gonadotropin / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D007700:Kinetics; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011989:Receptors, Thyrotropin; D013961:Thyroid Gland / Q000378:metabolism*; D013972:Thyrotropin / Q000378:metabolism*
chemical_list: D006063:Chorionic Gonadotropin; D011956:Receptors, Cell Surface; D011989:Receptors, Thyrotropin; D013972:Thyrotropin; D000262:Adenylyl Cyclases