Principles for Tuning Target Selectivity in Signaling Proteins

钙（Ca 2+）信号传导需要Ca 2+浓度随时间变化。钙调素（CaM）是一个主要的目标， 解码Ca 2+信号，但其固有的Ca 2+结合特性本身似乎不足以快速解码 Ca 2+信号波动。我们认为，除了下游信号转导，钙调素的目标， 通过构象调节介导的相互作用直接调节Ca 2+结合CaM的特性 这些调整增加了一种未发现的用于产生目标选择性的随时间变化的机制。此外，本发明还 我们提出，从ter-的角度来看，CaM的Ca 2+结合特性的靶诱导调谐， 分子反应是钙调素选择性是如何介导的拼图中缺失的一块。的目标 本研究旨在开发一种新的方法来表征CaM如何结合Ca 2+及其靶蛋白 这是CaM靶向选择性的核心特征。前几年完成的工作 资金期间表明，钙调素结合其目标是一个过程，涉及构象和相互 诱导的拟合和CaM和靶分子中的构象调整必须在 绑定的路径。我们的进展已经为这个持续项目的中心假设奠定了基础 Ca 2+结合环中的Ca 2+调节局部电场，并且环构象在 CaM的四只EF手这些EF手的整合动态调整了相互关系 在CaM的Ca 2+结合和靶结合之间，这对靶是独特的。我们进一步证明， 靶点选择性原理，具有两个拮抗剂靶点，神经颗粒蛋白和CaM依赖性激酶II， 与CaM相互作用，作为解码Ca 2+输入的不同模式的模块。理由是，一旦 确定了蛋白质靶点调节相互关系的原理，我们可以设计这样的蛋白质靶点 其控制CaM对不同频率或幅度的Ca 2+作出反应，用于在活细胞中转导信号。 我们将通过以下三个方面来检验我们的中心假设：（1） 钙调素与靶蛋白的反应不同，并在不同的条件下形成钙调素的整体构象。 Ca 2+结合的含量？(2)Ca ~（2+）如何调节其局部电场和EF的构象 对目标绑定的响应？(3)钙调素是如何从两个相互竞争的蛋白质靶点中进行选择的 对Ca 2+的亲和力相反的方向？这项研究是创新的，因为我们将开发一种新的 通过整合量子力学计算，分子计算， 模拟，生物物理和生物化学实验，以表征目标选择性如何可以 办妥了一批这项拟议的研究是有意义的，因为拟议的研究将产生一个 突破我们对这些过程的理解，并将提供洞察如何时间和 幅度变化的Ca 2+信号被CaM解码，并且CaM结合靶家族被协调成 生物反应。

Calcium (Ca2+) signaling requires Ca2+ concentration varying with time. Calmodulin (CaM) is a main target for decoding the Ca2+ signal but its intrinsic Ca2+-binding properties alone appear insufficient to decode rapidly fluctuating Ca2+ signals. We propose that in addition to transducing the signal downstream, CaM-targets directly tune the Ca2+-binding properties of CaM through reciprocal interactions mediated by conformational adjustments that add an undiscovered temporally varying mechanism for producing target selectivity. Further, we propose that the target induced tuning of CaM's Ca2+-binding properties from the perspective of ter- molecular reactions is the missing piece to the puzzle for how CaM selectivity is mediated. The objective of the present proposal is to develop a novel approach characterizing how CaM binds Ca2+ and its target protein reciprocally, which underlies the central feature of CaM's target selectivity. Work accomplished in the previous funding period has shown that CaM binding to its targets is a process involving conformationally and mutually induced fit and that the conformational adjustments in both CaM and target molecules must be overcome in the pathway of binding. Our progress has built the foundation for the central hypothesis in this continuing project that the Ca2+ in a Ca2+-binding loop tunes the local electric field and the loop conformations differentially among the four EF-hands of CaM. The integrative dynamics of these EF hands adjusts the reciprocal relations between CaM's Ca2+ binding and target binding, which is distinctive to a target. We further demonstrate the principle of target selectivity with a system of two antagonist targets, neurogranin and CaM-dependent kinase II, interacting with CaM as a module for decoding distinct patterns of Ca2+ input. The rationale is that once the principle of tuning the reciprocal relation by a protein target is identified, we can design such protein targets that control CaM to respond to different frequencies or amplitudes of Ca2+ for transducing signals in live cells. We will test our central hypothesis by pursuing the following three thrusts: (1) How does the four EF hands of CaM differentially react to a target protein and shape the global conformation of CaM under the variable content of Ca2+-binding? (2) How does Ca2+ tune its local electric field and conformations of an EF hand in response to target binding? (3) How does CaM select from the two competing protein targets that tune CaM's affinity for Ca2+ in opposite directions? The research proposed is innovative because we will develop a novel computer model for Ca2+ sensing in CaM by integrating quantum mechanical calculations, molecular simulations, and biophysical and biochemical experiments, to characterize how target selectivity can be achieved. The proposed research is of significance because the proposed study would engender a breakthrough in our understanding of these processes and will provide insights into how the time and amplitude varying Ca2+ signal is decoded by CaM and the family of CaM-binding targets into coordinated biological responses.