Cooperativity among calmodulin's drug binding sites.
Cooperativity among calmodulin's drug binding sites.
复制标题
钙调蛋白药物结合位点之间的协同作用。
DOI：
10.1021/bi00339a026
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发表时间：
1985
期刊：
影响因子：
2.9
通讯作者：
Johnson,JD
中科院分区：
文献类型：
--
作者：
Mills,JS;Bailey,BL;Johnson,JD
生理化学系，俄亥俄州州立大学医学中心，哥伦布，俄亥俄州43210接收1985年1月16日摘要：非洛地平，二氢吡啶钙离子拮抗剂，钙调蛋白的结合已被平衡透析和荧光技术研究。使用希尔方程分析得出希尔系数为2。结合态[非洛地平]与游离态[非洛地平] 2的曲线图显示Bmax为1.9 mol/mol，K 0 5为22 µ。两种钙调蛋白拮抗剂，异戊烯胺和R24571，先前已显示当非洛地平与钙调蛋白结合时可增强所观察到的荧光增强[约翰逊，J.D.（1983）生物化学。生物物理学。通信资源112，787]，分别使希尔系数降低至0.7和1.0，并解释了所观察到的非洛地平结合增强。在不存在和存在异戊烯胺和R24571的情况下，用钙调素滴定非洛地平表明，这些药物使非洛地平的钙调素的Kos降低25倍。因此，增效药物（异戊二烯胺和R24571）结合到非洛地平的两个结合位点中的任何一个，并通过变构机制，导致非洛地平以大大增强的亲和力结合到其余位点。观察到两种类型的增效药物。异戊烯胺显示希尔系数为0.8，而非洛地平、R24571和地尔硫卓在其增强非洛地平结合方面显示希尔系数为2。非洛地平和钙调素与Ca 2+的滴定表现出协同性，希尔系数为4。半数最大结合发生在pCa 6.0附近。在R24571存在下，非洛地平结合的钙依赖性是双相的，现在表现出更高的亲和力（pCa 7.6）组分。提出了一个模型来解释这些不同的变构调节构象的钙调素和它们的相互作用和激活与其靶蛋白的关系。（钙调素是一种普遍存在的钙结合蛋白，每摩尔蛋白质结合4摩尔钙，并经历大的钙依赖性结构变化。这些结构变化在其表面上形成或暴露疏水结合位点（Laporte等人，1980; Tanaka &日高，1980）其中cal-
Department of Physiological Chemistry, The Ohio State University Medical Center, Columbus, Ohio 43210 Received January 16, 1985 abstract: The binding of felodipine, a dihydropyridine Ca2+ antagonist, to calmodulin has been studied by equilibrium dialysis and fluorescence techniques. Analysis using the Hill equation gives a Hill coefficient of 2. A plot of bound [felodipine] vs. free [felodipine] 2 gives a Bmax of 1.9 mol/mol and a K0 5 of 22 µ. Two calmodulin antagonists, prenylamine and R24571, which have previously been shown to potentiate the fluorescent enhancement observed when felodipine binds to calmodulin [Johnson, J. D.(1983) Biochem. Biophys. Res. Commun. 112, 787], produce a reduction in Hill coefficient to 0.7 and 1.0, respectively, and account for the observed potentiation of felodipine binding. Titrations of felodipine with calmodulin in the absence and presence of prenylamine and R24571 suggest that these drugs decrease the Ko s of calmodulin for felodipine by 25-fold. Thus, potentiating drugs (prenylamine and R24571) bind to either of the two felodipine binding sites and, through an allosteric mechanism, result in felodipine binding to the remaining site with greatly enhanced affinity. Two types of potentiating drugs are observed. Prenylamine exhibits a Hill coefficient of 0.8 whereas felodipine, R24571, and diltiazem exhibit Hill coefficients of 2 in their potentiation of felodipine binding. Titrations of felodipine and calmodulin with Ca2+ exhibit cooperativity with a Hill coefficient of 4. Half-maximal binding occurs near pCa 6.0. In the presence of R24571, the calcium dependence of felodipine binding is biphasic, now exhibiting a much higher affinity (pCa 7.6) component. A model is presented to explain the relationship of these various allosterically regulated conformers of calmodulin and their interactions and activation with its target proteins.(Calmodulin is a ubiquitous calcium binding protein that binds 4 mol of calcium/mol of protein andundergoes large calcium-dependent changes in structure. These structural changes form or expose hydrophobic binding sites on its surface (Laporte et al., 1980; Tanaka & Hidaka, 1980) where cal-