Carayon P, Guibout M, Lissitzky S
Ann Endocrinol (Paris). 1979;40(3):211-27.
Plasma membranes have been purified from homogenates of normal human thyroid glands and multinodular euthyroid and Graves' goitres by discontinuous sucrose gradient centrifugation. Preparations of reasonable purity were obtained containing specific binding sites for thyrotropin and thyrotropin-sensitive adenylate cyclase. Optimum conditions for 125I-labeled thyrotropin binding were pH 7.8 and 37 degrees C. Sodium ions and concentration of Tris above 20 mM reduced thyrotropin binding. Human plasma membranes showed no species specificity toward thyrotropins from 3 different species (ox, hog and man). Displacement curves of I125-labeled bovine thyrotropin by unlabeled hormones was in the order of increasing concentrations of bovine, porcine and human highly purified thyrotropins and was inversely related to the specific biological activity of these preparations as determined by the bioassay in the mouse. Analysis of the interaction between membranes and 125I-labeled thyrotropin resulted in curvilinear Scatchard plots which can indicate the presence of two types of sites with high affinity -- low capacity (KD = 5 nM) and low affinity -- high capacity (KD = 500 nM) or site -- site interaction of the negative cooperativity type. No significant difference in binding site characteristics was found in normal and diseased glands (multinodular and Graves' goitres). A good correlation was found at equilibrium and in the conditions of adenylate cyclase assay between receptor occupancy and cyclase activation by b-thyrotropin.
通过不连续蔗糖梯度离心法，已从正常人甲状腺、多结节甲状腺功能正常者以及格雷夫斯病甲状腺肿的匀浆中纯化出质膜。获得了纯度合理的制剂，其中含有促甲状腺激素的特异性结合位点和对促甲状腺激素敏感的腺苷酸环化酶。¹²⁵I标记的促甲状腺激素结合的最佳条件是pH 7.8和37℃。钠离子和浓度高于20 mM的Tris会降低促甲状腺激素的结合。人质膜对来自3种不同物种（牛、猪和人）的促甲状腺激素没有物种特异性。未标记激素对¹²⁵I标记的牛促甲状腺激素的置换曲线，按照牛、猪和人高纯度促甲状腺激素浓度增加的顺序排列，并且与这些制剂通过小鼠生物测定法测定的特定生物活性呈负相关。对质膜与¹²⁵I标记的促甲状腺激素之间相互作用的分析产生了曲线型Scatchard图，这可能表明存在两种类型的位点，即高亲和力 - 低容量（KD = 5 nM）和低亲和力 - 高容量（KD = 500 nM），或者是负协同性类型的位点 - 位点相互作用。在正常和患病腺体（多结节和格雷夫斯病甲状腺肿）中，未发现结合位点特征有显著差异。在平衡状态下以及在腺苷酸环化酶测定条件下，发现β-促甲状腺激素的受体占有率与环化酶激活之间具有良好的相关性。
