Structure and stability of protein complexes in solution and the gas phase

大多数生物过程，包括免疫反应、细胞间通讯、炎症以及细菌和病毒感染，都涉及生物分子的缔合以形成特异性的非共价复合物。这些复合物的结构和稳定性由结合配偶体之间的许多力（例如氢键、离子和货车范德华相互作用）的协同作用以及与结合配偶体的溶剂壳相关的溶剂分子的置换和重组决定。理解这些力和它们导致的结构对于完整理解生物过程至关重要。 我们实验室的研究重点是基于质谱（MS）的技术的开发和应用，以在体外检测特定的非共价蛋白质复合物（例如蛋白质-配体和多蛋白质复合物），并表征其在溶液和气相中的结构和稳定性。该提案描述了新的实验方法的发展，基于电喷雾电离（ES）-MS技术，量化的动力学和热力学参数的非共价蛋白质在溶液中的相互作用。具体而言，我们将开发一种直接和灵敏的ES-MS方法来量化蛋白质与大分子（例如蛋白质和其他生物聚合物）相互作用的热力学参数。我们还将开发一种新的方法，基于温度跳跃松弛方法和ES-MS，以量化非共价蛋白质相互作用的动力学参数。从这些和其他方法的动力学和热力学数据将使我们能够开发一个更详细的图片的结构-功能的关系，在溶液中的蛋白质识别的基础。我们还将探讨使用红外多光子解离（IRMPD）光谱去溶剂化的非共价蛋白质复合物的结构。特别是，我们将评估这种技术的能力，探测气体蛋白质复合物内的分子间相互作用的性质。从比较的相互作用确定在溶液中的气相中，新的见解溶剂在生物识别中的作用将获得。

Most biological processes, including the immune response, cell-cell communication, inflammation and bacterial and viral infections, involve the association of biomolecules to form specific, non-covalent complexes. The structure and stability of these complexes are determined by the concerted action of many forces (e.g. hydrogen bonds, ionic and van der Waals interactions) between binding partners and from the displacement and reorganization of solvent molecules associated with the solvent shell of the binding partners. An understanding of these forces and the structures they lead to is essential to a complete understanding of biological processes. Research in our laboratory focuses on the development and application of mass spectrometry (MS)-based techniques to detect specific, non-covalent protein complexes (e.g. protein-ligand and multi-protein complexes) in vitro and to characterize their structure and stability in solution and the gas phase. This proposal describes the development of novel experimental methodologies, based on the electrospray ionization (ES)-MS technique, to quantify the kinetic and thermodynamic parameters for non-covalent protein interactions in solution. Specifically, we will develop a direct and sensitive ES-MS approach to quantify the thermodynamic parameters for protein interactions with macromolecules (e.g. proteins and other biopolymers). We will also develop a new method, based on the temperature-jump relaxation approach and ES-MS, to quantify the kinetic parameters for non-covalent protein interactions. The kinetic and thermodynamic data available from these and other methods will allow us to develop a more detailed picture of the structure-function relationships that underlie protein recognition in solution. We will also explore the structure of desolvated non-covalent protein complexes using infrared multiphoton dissociation (IRMPD) spectroscopy. In particular, we will assess the ability of this technique to probe the nature of intermolecular interactions within the gaseous protein complexes. From a comparison of the interactions identified in solution in the gas phase, new insights into the role of solvent in biological recognition will be gained.