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KINETIC AND BIOPHYSICAL ANALYSIS OF THE M2 MUSCARINIC RECEPTOR

机译:M2肌肉受体的运动学和生物物理分析

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The recombinant Pm2 muscarinic receptor expressed in Chinese hamster ovary (CHO) cells was used as a model system to examine receptor-effector coupling and ligand binding. In CHO cells, equilibrium binding studies and the dependence on receptor number per cell of the maximum response and EC(50) values for agonist stimulation of phosphatidylinositol metabolism and inhibition of cAMP formation were consistent with a modified ternary complex model of signal transduction that included a physiologically noncompetent receptor state. Detailed kinetic studies of oxotremorine M (Oxo-M) binding to CHO cell membranes suggested that agonist interactions at the high affinity class of binding sites are complicated and depend on receptor expression levels. At low levels of expression, kinetic data were consistent with a special case of a mechanism in which Oxo-M shifts the equilibrium between two receptor conformations while at high levels of expression, it was necessary to evoke receptor-receptor interactions to explain the kinetic data. Far ultraviolet circular dichroism studies of the purified recombinant receptor showed a high content of alpha-helical secondary structure and small changes in secondary structure upon antagonist, but not agonist, binding. [References: 11]
机译:在中国仓鼠卵巢(CHO)细胞中表达的重组Pm2毒蕈碱受体用作模型系统,以研究受体-效应子偶联和配体结合。在CHO细胞中,平衡结合研究以及对每个细胞受体数量的依赖性(最大响应和EC(50)值)对激动剂刺激磷脂酰肌醇代谢和cAMP形成的抑制作用与经修饰的信号传导三元复合模型相一致,该模型包括生理上不称职的受体状态。 oxotremorine M(Oxo-M)与CHO细胞膜结合的详细动力学研究表明,结合位点的高亲和力类的激动剂相互作用很复杂,并且取决于受体的表达水平。在低水平表达时,动力学数据与其中Oxo-M改变两个受体构象之间的平衡的机制的特殊情况一致,而在高水平表达时,有必要引起受体-受体相互作用来解释动力学数据。纯化的重组受体的远紫外圆二色性研究表明,α-螺旋二级结构的含量很高,而拮抗剂结合却没有激动剂,二级结构的变化很小。 [参考:11]

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