FORCE-MODULATED BINDING AFFINITY: COMPUTATIONAL STUDY OF FAT-PAXILLIN INTERACTI

这个子项目是许多利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 力学信号可以调节细胞的生长、分化、凋亡、运动和基因表达等多种生理行为。机械信号导致生物化学和细胞变化或机械转导的过程的细节在很大程度上是未知的。一种提出的机制是力驱动的构象变化导致结合亲和力改变，基本上将机械信号转化为浓度变化。我们将通过对蛋白质拉动模拟的详细结构和能量分析来表征力引起的结合变化的特性。方法上的工作使用了多维副本交换协议，通过更好的采样加快收敛，和加权直方图分析方法（WHAM）计算自由能的变化引起的机械扰动。我们的工作重点是粘着斑激酶与桩蛋白肽结合的粘着斑靶向（FAT）结构域。粘着斑是细胞指向细胞外环境的锚点，是已知作为机械感觉装置的动态蛋白质复合物，其中机械力可以调节位点的组装并触发信号传导。虽然响应于力的蛋白质的精确身份尚未阐明，但FAT和桩蛋白之间的相互作用在粘着斑形成中起重要作用，并且被认为是机械传感机制的一部分。初步模拟表明，力可以通过激活新的接触来诱导加强脂肪-桩蛋白结合相互作用。我们计划调查变量效应沿着不同的拉动方向，以深入了解这种反应在生物学背景下的鲁棒性，以及进行突变研究的受力残基。详细了解力学转导的分子机制可能会导致治疗和组织工程的进步，设计模拟细胞天然化学和机械环境的培养条件。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Mechanical signals have been shown to regulate various physiological behaviors, including cell growth, differentiation, apoptosis, motility and gene expression. The details of the processes by which mechanical cues result in biochemical and cellular change, or mechanotransduction, are largely unknown. One proposed mechanism is that a force-driven conformational change results in altered binding affinity, essentially converting a mechanical signal into a concentration change. We will characterize properties of force-induced binding changes, through detailed structural and energetic analysis of protein pulling simulations. Methodologically the work uses a multi-dimensional replica exchange protocol, which speeds up convergence through better sampling, and the Weighted Histogram Analysis Method (WHAM) to compute free energy changes induced by mechanical perturbation. We are focusing our efforts on the focal adhesion targeting (FAT) domain of focal adhesion kinase binding to a paxillin peptide. Focal adhesions, the cell anchor points to the extracellular environment, are dynamical protein complexes known to act as mechanosensory devices, where mechanical forces can regulate the assembly of the site and trigger signaling. While the precise identity of proteins responsive to force is not elucidated, interactions between FAT and paxillin play an important role in focal adhesion formation and are thought to be part of the mechanosensing machinery. Preliminary simulations have shown that force can induce strengthening of FAT-paxillin binding interactions through activation of new contacts. We plan to investigate variable effects along different pulling directions to gain insight into the robustness of this response in a biological context, as well as to carry out mutational studies of force-bearing residues. Detailed understanding of the molecular mechanisms of mechanotransduction could lead to advances in therapeutics and tissue engineering, with design of culture conditions mimicking the cells natural chemical and mechanical environment.