Mode of coupling between the beta-adrenergic receptor and adenylate cyclase in turkey erythrocytes.
Mode of coupling between the beta-adrenergic receptor and adenylate cyclase in turkey erythrocytes.
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火鸡红细胞中β-肾上腺素能受体和腺苷酸环化酶之间的偶联模式。
作者：
A. Tolkovsky;A. Levitzki
详细分析了β -肾上腺素能受体与火鸡红细胞膜腺苷酸环化酶的偶联方式。已经使用了许多实验技术：(1)测量环化酶在胍基酰亚胺二磷酸存在下的永久活性状态的动力学，作为激素浓度的函数；(2)激素和胍基酰亚胺二磷酸激活酶之前和之后β -肾上腺素能受体拮抗剂和拮抗剂结合的测定。在这两种方法的基础上，所有涉及酶与受体之间平衡的受体与酶偶联模型都可以被拒绝。然而，结合和动力学数据可以用两种截然相反的受体与酶偶联模型来拟合：(a)酶激活的预偶联酶-受体模型，根据以下方案：公式（见原文）其中H是激素，RE是预偶联的受体-酶复合物，k1和k2是描述激素结合的速率常数，k是表征中间HRE形成HRE'的速率常数。根据这个模型，活化的复合物是由所有相互作用的物质组成的。(b)另一个模型是碰撞耦合机制：公式（见测试），其中KH为激素受体解离常数，k1为控制HRE形成的双分子速率常数，k3为控制酶活化的速率常数。在这种情况下，中间体不会积累，只占总受体和腺苷酸环化酶浓度的一小部分。为了确定这两种机制中哪一种支配腺苷酸环化酶受体激活模式，我们进行了一项诊断实验：发现通过特定亲和力标签逐渐使β受体失活，导致受体的最大结合能力下降，激活率成比例下降，但最大活性水平没有变化。对羟基汞苯甲酸酯使酶逐渐失活并不会改变酶的活化速率，也不会改变受体结合激素的能力。只有最大激活水平被发现降低。这些结果不符合受体和环化酶的预耦联模型，也不符合浮动受体模型，其中激素，受体和环化酶的中间体与其反应物处于平衡状态。这些数据强烈提示碰撞偶联是火鸡红细胞膜β受体和环化酶偶联的偶联方式。
The mode of coupling of the beta-adrenergic receptor to the enzyme adenylate cyclase in turkey erythrocyte membranes was analyzed in detail. A number of experimental techniques have been used: (1) measurement of the kinetics of cyclase activation to its permanetly active state in the presence of guanylyl imidodiphosphate, as a function of hormone concentrations; (2) measurement of antagonist and agoinst binding to the beta-adrenergic receptor prior and subsequent to the enzyme activation by hormone and guanylyl imidodiphosphate. On the bases of these two approaches, all the models of receptor to enzyme coupling which involve an equilibrium between the enzyme and the receptor can be rejected. The binding and the kinetic data, however, can be fitted by two diametrically opposed models of receptor to enzyme coupling: (a) the precouped enzyme-receptor model where activation of the enzyme occurs, according to the following scheme: formula (see text) where H is the hormone, RE is the precoupled respetor-enzyme complex, k1 and k2 are the rate constants describing hormone binding, and k is the rate constant characterizing the formation of HRE' from the intermediate HRE. According to this model, the activated complex is composed of all of the interacting species. (b) The other model is the collision coupling mechanism: formula (see test) wheere KH is the horome-receptor dissociation constant, k1 is the bimolecular rate constant governing the formation of HRE, and k3 the rate constant governing the activation of the enzyme. In this case the intermediate never accumulates and constitutes only a small fraction of the total receptor and adenylate cyclase concentrations. In order to establish which of the two mechanisms governs the mode of adenylate cyclase activation by its receptor, a diagnostic experiment was performed: Progressive inactivation of the beta receptor by a specific affinity label was found to cause a decrease in the maximal binding capacity of the receptor and a proportional decrease in the rate of activation, but no change in the maximum level of activity was attained. Progressive inactivation of the enzyme by p-hydroxymercuribenzoate was found not to change the rate of activation nor the capacity of the receptor to bind hormone. Only the maximal level of activation was found to be decreased. These results are not compatible with the precoupled model of receptor and cyclase nor with floating receptor models in which an intermediate of hormone, receptor, and cyclase is in equilibrium with its reactants. The data strongly suggest that the collision coupling is the mode of coupling between the beta receptor and cyclase coupling in turkey erythrocyte membranes.