Protein structure and dynamics in ultra-heterogeneous environments

超异质环境中的蛋白质结构和动力学

• 批准号: 10408147
• 负责人: Carlos Raul Baiz
• 金额: $ 20.58万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10408147
• 关键词: Behavior; Binding; Biological Process; Biophysics; Buffers; Calmodulin; Cardiovascular Diseases; Cells; Complex; Crowding; Cytoplasm; Disease; Electrostatics; Environment; Heterogeneity; Hydrogen Bonding; Ion Channel; Ion Channel Gating; Measurement; Mediating; Modeling; Molecular; Mutation; Physiological; Plant Roots; Protein Dynamics; Proteins; Role; Structure; System; Testing; Thermodynamics; Translating; Tube; Vertebral column; Water; in vivo; nervous system disorder; patch clamp; peptidomimetics; protein protein interaction; protein structure

SUMMARY Crowding and heterogeneity: Biomolecular organization in vivo is driven by crowding and heterogeneity. To date, protein structure, dynamics, and folding have been studied almost exclusively in simple buffer solutions, yet it is has recently become evident that most “test tube” studies cannot be directly translated to cellular environments. Nonspecific electrostatic interactions, excluded volume effects, and disrupted hydrogen-bond networks dictate protein thermodynamics in these complex environments. While the prevailing view from these is that excluded-volume effects favor the more compact native states, our group, along with others, found that enthalpic contributions strengthen protein-water hydrogen bonds. These interactions can increase backbone exposure and consequently destabilize folded states. Thus, there is an immediate need to quantify interactions between biomolecules in accurate cell-like environments. The present studies are critical first step towards understanding protein structure and dynamics in vivo. Our project aims to characterize the structure, dynamics, and stability of proteins in crowded solutions that accurately mimic the cytoplasm. Specifically, we will quantify the degree of molecular heterogeneity and establish the role of macromolecular crowding on protein-protein and protein-water contacts. Protein-protein interactions and ion channel gating mechanisms: Calmodulin (CaM) regulates biological function by modulating the behavior of a wide range of proteins including many ion channels. CaM mutations or mutations within CaM-regulated ion channels are responsible for neurological and cardiovascular diseases. CaM can be considered a “Ca-sensing domain” for multiple ion channels, but the dynamic association between CaM and ion channels make mechanistic studies challenging. The first complete structures of an ion channel with CaM were solved earlier this year (2018). These underscore the fact that the gating mechanisms remain incompletely understood. For example, eight states are required to model patch clamp measurements, but only two structures (open/closed) are known. We propose to investigate gating mechanisms through a detailed biophysical examination of dynamic CaM-channel interactions using a peptide that mimics the CaM binding domain of the SK2 channel (KCa2.2). SK channels are important in a wide variety of physiological systems and offer many advantages as a system for understanding Ca2+-CaM-mediated gating. If successful, our studies will produce a stepwise mechanistic view of CaM-mediated channel activation.

概括 拥挤与异质性：体内生物分子组织由拥挤和异质性驱动。到 日期，蛋白质结构，动力学和折叠几乎完全是在简单的缓冲溶液中研究的， 然而，最近已经成为证据，表明大多数“试管”研究不能直接翻译成细胞 环境。非特异性静电相互作用，排除体积效应和氢键中断 网络在这些复杂环境中决定蛋白质热力学。而这些景观 是排除的体积效应有利于更紧凑的本土国家，我们的群体以及其他人发现 焓贡献增强了蛋白质 - 水氢键。这些相互作用会增加主干 暴露并因此破坏了折叠状态。这是立即量化互动的需求 在精确细胞样环境中的生物分子之间。目前的研究是迈向的关键第一步 了解体内蛋白质结构和动力学。我们的项目旨在表征结构，动态， 蛋白质在拥挤的溶液中的稳定性，这些溶液准确地模仿了细胞质。具体而言，我们将量化 分子异质性的程度，并确定大分子拥挤在蛋白质蛋白质和 蛋白质水接触。 蛋白质 - 蛋白质相互作用和离子通道门控机制：钙调蛋白（CAM）调节生物学 通过调节包括许多离子通道在内的广泛蛋白质的行为来调节功能。凸轮突变或 凸轮调节的离子通道内的突变导致神经系统疾病和心血管疾病。凸轮 可以认为是多个离子通道的“ CA敏感域”，但是CAM之间的动态关联 离子渠道使机械研究挑战。离子通道的第一个完整结构， CAM在今年早些时候（2018年）得到了解决。这些强调了门控机制仍然存在的事实 不完全理解。例如，需要八个状态才能建模斑块夹的测量，但仅需要 已知两个结构（开放/关闭）。我们建议通过详细的 使用模拟凸轮结合的肽对动态凸轮通道相互作用的生物物理检查 SK2通道的域（KCA2.2）。 SK渠道在各种物理系统中都很重要， 提供许多优势作为了解CA2+CAM介导的门控的系统。如果成功，我们的学习将 产生CAM介导的通道激活的逐步机械视图。