Dynamic Properties of Bacterial Adhesins

描述(由申请人提供)：在这项竞争性的更新申请中提出的研究旨在剖析拉伸机械力如何提高一些最常见的细菌粘附素与碳水化合物配体结合的能力，即介导受体-配体相互作用的捕获键机制。我们目前的生物工程研究伙伴资助项目已经证明，剪切应力的存在大大增强了甘露糖特异性的、FimH介导的大肠杆菌的粘附力。我们已经证明FimH粘附素是一种变构调控的蛋白质，其中甘露糖配体与FimH结合口袋相互作用的诱导适配机制与FimH的甘露糖结合结构域和菌毛结合结构域之间的分离在构象上相连--这种构型有利于拉伸机械力的作用。在更新应用中，我们建议使用核磁共振(NMR)光谱和原子力显微镜(AFM)以及分子动力学模拟(MD/SMD)等方法来全面了解FimH在配体激活过程中的构象变化，以及通过张力力促进这一过程。我们将利用这些知识来制定防止医学相关细菌与宿主靶细胞和表面黏附的策略，开发剪切调制纳米技术工具，并将其作为理解其他类型的剪切依赖细菌黏附的范例。

DESCRIPTION (provided by applicant): Studies proposed in this competing renewal application are intended to dissect how tensile mechanical force increases ability of some of the most common bacterial adhesins to bind carbohydrate ligands, i.e. mediate catch-bond mechanism of receptor-ligand interaction. Our current Bioengineering Research Partnership grant project has demonstrated that the strength of mannose-specific, FimH-mediated adhesion of Escherichia coli is dramatically enhanced by the presence of shear stress. We have shown that FimH adhesin is an allosterically regulated protein, where induced-fit mechanism of interaction of the mannose ligand with the FimH binding pocket is conformationally linked with separation between mannose-binding and fimbria-incorporating domains of FimH - the configuration favored by tensile mechanical force. In the renewal application, we propose to use, among other approaches, nuclear magnetic resonance (NMR) spectroscopy and atomic force microscopy (AFM) and molecular dynamics simulation (MD/SMD) to derive a comprehensive understanding of the conformational shift in course of FimH activation by the ligand and facilitation of this process by tensile force. We will use this knowledge for developing strategies on preventing adhesion of medically-relevant bacteria to host target cells and surfaces, for developing shear- modulated nanotechnological tools, and as a paradigm for understanding other types of shear-dependent bacterial adhesion.