Understanding membrane proteins' allosteric modulation with cryo-EM

利用冷冻电镜了解膜蛋白的变构调节

• 批准号: 11002891
• 负责人: Amedee des Georges
• 金额: $ 15.38万
• 依托单位: NEW YORK UNIVERSITY
• 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 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/11002891
• 关键词: Understanding; membrane; proteins; allosteric; modulation

Project Summary/Abstract My laboratory's research is focused on understanding the molecular mechanisms key to the regulation of membrane protein function and to the modulation of their output signaling. We use cryo-EM and advanced classification methods, such as manifold embedding, combined with modeling and molecular dynamics to study how ligands produce shifts in conformation equilibria and bias signaling output. We are currently studying two systems that allow us to study how different types of ligands influence the gating of ion channels. We are studying the role of lipids in triggering gating of mechanosensitive channels of the MscS family, a model system of membrane tension-sensing which can yield important information about the fundamental principles governing ion channel gating and the role that lipid-protein interactions play in this process. We have a longstanding interest in the mechanism and modulation of the ryanodine receptor (RyR), a calcium release channel fundamental to heart and skeletal function. We are using manifold embedding, a machine learning-based method to analyze cryo-EM images, to better understand how small molecules and ions such as calcium and ATP affect the conformational energy landscape of the channel. We aim to shed light on the gating mechanism of this very large ion channel, a prerequisite to understanding of its modulation by protein ligands, post-translational modifications and drugs. Our approach to these fundamental problems is to use a commbination of cryo-electron microscopy, advanced image classification techniques, modelling and molecular dynamics simulations to delineate allosteric pathways mechanistically and then test proposed models with biophysical methods such as single-channel measurements, HDX-MS and mutagenesis. Our ultimate aim is to greatly increase our molecular understanding of the gating and allosteric modulation of ion channels. Progress towards such knowledge has the potential to open the way for the design of small molecule allosteric modulators with very well controlled effects on their targets, aiding the development of drugs with limited side effects. A longer-term aim is to further develp and use our tools towards gaining a better understanding of allosteric modulation in other classes of membrane proteins of therapeutic importance. In particular, G protein-coupled receptor ligands can induce the selective binding of different transducers in a process called biased signaling. The molecular mechanisms at play in this process are still poorly understood despite their fundamental importance for the development of safer drugs.

项目概要/摘要 我实验室的研究重点是了解调节的关键分子机制 膜蛋白功能及其输出信号的调节。我们使用冷冻电镜和先进的 分类方法，例如流形嵌入，与建模和分子动力学相结合 研究配体如何产生构象平衡的变化和偏差信号输出。我们目前正在学习 两个系统使我们能够研究不同类型的配体如何影响离子通道的门控。 我们正在研究脂质在触发 MScS 家族机械敏感通道门控中的作用，该家族是一个 膜张力传感模型系统，可以产生有关基本原理的重要信息 控制离子通道门控的原理以及脂质-蛋白质相互作用在此过程中发挥的作用。 我们对兰尼碱受体 (RyR) 的机制和调节有着长期的兴趣，兰尼碱受体是一种钙 释放通道对心脏和骨骼功能至关重要。我们正在使用流形嵌入，一台机器 基于学习的方法来分析冷冻电镜图像，以更好地了解小分子和离子等 因为钙和 ATP 会影响通道的构象能量景观。我们的目标是阐明 这个非常大的离子通道的门控机制，是理解蛋白质调节的先决条件 配体、翻译后修饰和药物。 我们解决这些基本问题的方法是结合使用冷冻电子显微镜、先进的 图像分类技术、建模和分子动力学模拟来描绘变构 机械地分析路径，然后使用生物物理方法（例如单通道）测试所提出的模型 测量、HDX-MS 和诱变。 我们的最终目标是大大增加我们对门控和变构调节的分子理解 离子通道。这些知识的进展有可能为小型设计开辟道路。 分子变构调节剂对其靶标具有很好的控制效果，有助于开发 副作用有限的药物。 长期目标是进一步开发和使用我们的工具来更好地了解变构 对具有治疗重要性的其他类别膜蛋白的调节。特别是，G蛋白偶联 受体配体可以在称为偏向信号传导的过程中诱导不同传感器的选择性结合。 尽管具有基本原理，但在此过程中发挥作用的分子机制仍然知之甚少。 对于开发更安全的药物具有重要意义。

项目成果

Visualization of translation reorganization upon persistent ribosome collision stress in mammalian cells.

哺乳动物细胞中持续核糖体碰撞应激下翻译重组的可视化。

• DOI: 10.1016/j.molcel.2024.01.015
• 发表时间: 2024
• 期刊: Molecular cell
• 影响因子: 16
• 作者: Fedry,Juliette;Silva,Joana;Vanevic,Mihajlo;Fronik,Stanley;Mechulam,Yves;Schmitt,Emmanuelle;desGeorges,Amédée;Faller,WilliamJames;Förster,Friedrich
• 通讯作者: Förster,Friedrich