Assessing the Role of Calmodulin's Linker Flexibility in Target Binding.
Assessing the Role of Calmodulin's Linker Flexibility in Target Binding.
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评估钙调蛋白的接头柔性在靶结合中的作用。
DOI：
10.3390/ijms22094990
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发表时间：
2021-05-08
影响因子：
5.6
通讯作者：
Kekenes-Huskey PM
中科院分区：
文献类型：
--
作者：
Sun B;Kekenes-Huskey PM
钙调素（CaM）是一种高表达的钙结合蛋白，已知可结合数百种蛋白质靶点。其对许多这些靶标的结合选择性部分归因于蛋白质的连接其N-和C-结构域的柔性α螺旋接头。目前还没有很好地确定它的接头如何介导钙调素的结合到监管目标。深入了解这一点将是非常宝贵的，以了解其调节不同的细胞信号通路。因此，我们利用马提尼粗粒度（CG）分子动力学模拟探测钙调素/目标组件的模型系统：钙调素结合钙调神经磷酸酶（CaN）的监管结构域。进行模拟，假设“野生型”钙调蛋白具有其接头的正常柔性，以及不稳定的、高度柔性的接头变体，以模拟例如可以通过翻译后修饰诱导的结构变化。对于野生型模型，在三种离子强度的600次模拟中，有98%在2 μs的模拟时间内采用了结合复合物;其中，1.7%采样了在实验确定的晶体结构中观察到的完全结合状态。通过计算这些模拟的平均第一时间，我们估计缔合速率为8.7 × 10 M s，这与实验确定的扩散限制速率2.2 × 10 M s相似。此外，我们的模拟概括了其众所周知的缔合速率和溶液离子强度之间的反比关系。相比之下，虽然超过97%的不稳定接头模拟形成紧密结合的复合物，但只有0.3%实现了完全结合的构型。这种效应似乎源于由连接体柔性控制的扩展和塌陷状态的集合的差异。因此，我们的模拟表明，在钙调素接头的α螺旋二级结构的倾向的变化可以调节靶结合的动力学。
Calmodulin (CaM) is a highly-expressed Ca binding protein known to bind hundreds of protein targets. Its binding selectivity to many of these targets is partially attributed to the protein’s flexible alpha helical linker that connects its N- and C-domains. It is not well established how its linker mediates CaM’s binding to regulatory targets yet. Insights into this would be invaluable to understanding its regulation of diverse cellular signaling pathways. Therefore, we utilized Martini coarse-grained (CG) molecular dynamics simulations to probe CaM/target assembly for a model system: CaM binding to the calcineurin (CaN) regulatory domain. The simulations were conducted assuming a ‘wild-type’ calmodulin with normal flexibility of its linker, as well as a labile, highly-flexible linker variant to emulate structural changes that could be induced, for instance, by post-translational modifications. For the wild-type model, 98% of the 600 simulations across three ionic strengths adopted a bound complex within 2 μs of simulation time; of these, 1.7% sampled the fully-bound state observed in the experimentally-determined crystallographic structure. By calculating the mean-first-passage-time for these simulations, we estimated the association rate to be 8.7 × 10 M s, which is similar to the diffusion-limited, experimentally-determined rate of 2.2 × 10 M s. Furthermore, our simulations recapitulated its well-known inverse relationship between the association rate and the solution ionic strength. In contrast, although over 97% of the labile linker simulations formed tightly-bound complexes, only 0.3% achieved the fully-bound configuration. This effect appears to stem from a difference in the ensembles of extended and collapsed states which are controlled by the linker flexibility. Therefore, our simulations suggest that variations in the CaM linker’s propensity for alpha helical secondary structure can modulate the kinetics of target binding.
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