Interactions of membrane proteins with lipids, water, and other proteins in biological membranes

膜蛋白与生物膜中的脂质、水和其他蛋白质的相互作用

• 批准号: RGPIN-2014-04547
• 负责人: Ladizhansky, Vladimir
• 金额: $ 3.86万
• 依托单位: University of Guelph
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2014
• 资助国家: 加拿大
• 起止时间: 2014-01-01 至 2015-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=566590
• 关键词: Interactions; membrane; proteins; lipids; water

BACKGROUND Membrane proteins constitute approximately a third of all proteins. They are involved in many important physiological processes, and represent almost a half of known drug targets. Studying membrane proteins in their native lipid bilayer environment is particularly challenging, as the two common structural methods, X-ray crystallography and solution-state Nuclear Magnetic Resonance (NMR), rely on crystals or detergent micelles, which are different from that of lipids. Solid-state NMR (SSNMR) provides an alternative approach to protein structure determination, and offers ways to study membrane proteins in the bilayer environment. OBJECTIVES In this application, we will pursue both the methodological objectives aimed at improving SSNMR ability to characterize protein structures and protein interactions, and to answer the fundamental biophysical questions related to the response of membrane protein conformations to changes in the environment. Our goal is to undestand how interactions between membrane proteins and their environment shape protein structure. Specifically, we will focuss on the following three types of effects: (i) Protein-water interactions. Water molecules play important roles in defining the structure, stability, dynamics and function of proteins. Buried waters are extensively hydrogen-bonded with backbone and side chain atoms, both stabilizing the secondary and tertiary structures, and playing functional roles. As protein-water interactions are inherently dynamic, NMR is ideally suited to study these interactions. We are interested in determining the location of bound water molecules, and in characterizing their dynamics. (ii) Protein-lipid interactions. We are interested in characterizing the effects of both specific and non-specific interactions, e.g., how tightly bound non-annular lipids affect the protein structure, and how variation of the bilayer properties affect membrane proteins structure, oligomerization and dynamics. (iii) Structure of membrane proteins in cellular membranes. Studying biomolecules in the cellular environment is the ultimate goal of molecular biology. SSNMR provides an excellent tool to elucidate the structural and dynamic changes in membrane proteins induced by the complex environment of E. coli membranes with many additional endogenous molecular components present. The fundamental question we strive to answer is to what extent the idealized environment of a reconstituted bilayer resembles that of a real membrane. SIGNIFICANCE A growing number of SSNMR structures of lipid-embedded membrane proteins underscores the potential of solid-state NMR as a powerful method to investigate membrane proteins in the native-like bilayer environment. The present proposal aims to go beyond the structure and to study functionally and structurally important interactions between membrane proteins, and water and lipids. We will develop new SSNMR methodologies for obtaining comprehensive view of membrane protein structure and dynamics that include bound water and lipid molecules, and will contribute detailed structural information on the long-standing questions of the effects of environment on the membrane protein functional assembly, structure, and dynamics.

膜蛋白约占所有蛋白质的三分之一。它们参与许多重要的生理过程，并且代表了几乎一半的已知药物靶标。在天然脂质双层环境中研究膜蛋白特别具有挑战性，因为两种常见的结构方法，X射线晶体学和溶液状态核磁共振（NMR），依赖于晶体或洗涤剂胶束，这与脂质不同。固态核磁共振（SSNMR）提供了一种替代方法来确定蛋白质结构，并提供了在双层环境中研究膜蛋白的方法。在这个应用程序中，我们将追求两个方法的目标，旨在提高SSNMR的能力，表征蛋白质结构和蛋白质相互作用，并回答有关的膜蛋白构象的响应在环境中的变化的基本生物物理问题。我们的目标是了解膜蛋白与其环境之间的相互作用如何塑造蛋白质结构。具体而言，我们将重点关注以下三种类型的影响：（i）蛋白质-水相互作用。水分子在定义蛋白质的结构、稳定性、动力学和功能方面起着重要作用。埋藏的沃茨与主链和侧链原子广泛氢键结合，既稳定二级和三级结构，又发挥功能作用。由于蛋白质与水的相互作用具有内在的动态性，因此NMR非常适合研究这些相互作用。我们感兴趣的是确定结合水分子的位置，并在其动态特性。(ii)蛋白质-脂质相互作用。我们感兴趣的是表征特异性和非特异性相互作用的影响，例如，紧密结合的非环状脂质如何影响蛋白质结构，以及双层性质的变化如何影响膜蛋白质结构、寡聚化和动力学。(iii)细胞膜中膜蛋白的结构。研究细胞环境中的生物分子是分子生物学的最终目标。SSNMR为阐明E.大肠杆菌膜与许多额外的内源性分子成分存在。我们努力回答的基本问题是，重建双层的理想环境在多大程度上类似于真实的膜。越来越多的脂质包埋膜蛋白的SSNMR结构强调了固态NMR作为一种强有力的方法来研究膜蛋白在天然样双层环境中的潜力。本提案旨在超越结构，并研究膜蛋白，水和脂质之间的功能和结构上的重要相互作用。我们将开发新的SSNMR方法，用于获得膜蛋白结构和动力学的全面视图，包括结合水和脂质分子，并将有助于详细的结构信息的长期存在的问题，环境对膜蛋白功能组装，结构和动力学的影响。