Mapping membrane protein dynamics in time and space with mass spectrometry

利用质谱绘制膜蛋白的时间和空间动态图

• 批准号: EP/V011715/1
• 负责人: Argyris Politis
• 金额: $ 129.78万
• 依托单位: University of Manchester
• 依托单位国家: 英国
• 项目类别: Fellowship
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=EP%2FV011715%2F1
• 关键词: Mapping; membrane; protein; dynamics; time

The advancement of new analytical tools and methods are at the centre of tackling the biggest challenges in the life sciences. One such unmet challenge is understanding the structural dynamics underpinning function in membrane-embedded proteins. Membrane proteins control what comes in and what goes out of the cell. As a consequence, they constitute the main targets of more than half of known drugs. Despite their critical importance, existing methods often fail to uncover structural information about this important class of biomolecules, thus precluding progresses related to therapeutic intervention and drug discovery. More importantly, we currently lack the tools to capture the dynamics of membrane proteins within their native environment wherein they move and operate. This is primarily due to the complexity of such systems as they are embedded into a heterogeneous layer of lipids, which protect their hydrophobic core of membrane proteins. New tools are therefore urgently needed to unveil the dynamic motions of membrane proteins and allow mechanistic insights important for addressing current and future challenges related to human health and disease.Here, we will built a new method to capture molecular movies of membrane proteins in action. To do this, we will develop time-resolved hydrogen deuterium exchange mass spectrometry (tHDX-MS). HDX-MS is a sensitive analytical tool that can accurately monitor the exchange of hydrogen atoms with the heavier deuterium in solution, thus offering information about protein dynamics. By combining the emerging HDX-MS technology with microfluidic techniques, we will enable snapshots of membrane protein states in times ranging from microseconds to hours and with adjustable resolution. To enable applicability of our strategy within the native lipid environment wherein membrane proteins function, we will utilise the controlled patches of membrane biomimetics, the so called nanodiscs. The nanodisc technology will allow us to fine-tune the lipid composition surrounding membrane proteins and assess the individual effect of specific lipids on membrane protein structure and dynamics. We will demonstrate applicability of our approach on a range of important systems of increasing size and complexity including the challenging G protein-coupled receptors (GPCRs) that are the key drug targets. To make our approach amenable to large and dynamic complexes and circumvent current challenges with respect to sensitivity and resolution, we will work with our industrial partner (Waters Corp.) to utilise a currently non-commercial, prototypical instrumentation (Cyclic HDX-MS). This together with our methodological advancements will allow us to be the first to achieve this for membrane proteins in lipid context and thus become the leaders in this rapidly evolving field of research in the UK and worldwide. Overall, this fellowship will not only establish a new tool for tackling key challenges in deciphering the dynamic mechanisms underpinning membrane protein function but it will also allow me to lead this exciting and emerging field of research, currently under-represented in the UK.

新分析工具和方法的进步是应对生命科学中最大挑战的核心。其中一项未解决的挑战是了解膜嵌入蛋白功能的结构动力学基础。膜蛋白控制细胞的进出。因此，它们构成了一半以上已知药物的主要靶标。尽管它们至关重要，但现有方法往往无法揭示这一类重要生物分子的结构信息，从而阻碍了与治疗干预和药物发现相关的进展。更重要的是，我们目前缺乏捕获膜蛋白在其移动和运行的原生环境中动态的工具。这主要是由于此类系统的复杂性，因为它们嵌入到异质脂质层中，从而保护其膜蛋白的疏水核心。因此，迫切需要新的工具来揭示膜蛋白的动态运动，并为解决当前和未来与人类健康和疾病相关的挑战提供重要的机制见解。在这里，我们将建立一种新方法来捕获膜蛋白运动的分子电影。为此，我们将开发时间分辨氢氘交换质谱 (tHDX-MS)。 HDX-MS 是一种灵敏的分析工具，可以准确监测溶液中氢原子与较重氘的交换，从而提供有关蛋白质动力学的信息。通过将新兴的 HDX-MS 技术与微流体技术相结合，我们将能够在微秒到几小时的时间内以可调节的分辨率拍摄膜蛋白状态的快照。为了使我们的策略在膜蛋白发挥作用的天然脂质环境中适用，我们将利用膜仿生学的受控斑块，即所谓的纳米圆盘。纳米圆盘技术将使我们能够微调膜蛋白周围的脂质成分，并评估特定脂质对膜蛋白结构和动力学的个体影响。我们将证明我们的方法对一系列规模不断增加和复杂性不断增加的重要系统的适用性，包括作为关键药物靶标的具有挑战性的 G 蛋白偶联受体 (GPCR)。为了使我们的方法能够适应大型动态复合物并规避当前在灵敏度和分辨率方面的挑战，我们将与我们的工业合作伙伴（沃特世公司）合作，利用目前非商业化的原型仪器（循环 HDX-MS）。这与我们方法论的进步一起将使我们成为第一个在脂质背景下实现膜蛋白这一目标的公司，从而成为英国和全球这一快速发展的研究领域的领导者。总的来说，这项奖学金不仅将建立一个新工具来应对破译支撑膜蛋白功能的动态机制的关键挑战，而且还将使我能够领导这个令人兴奋的新兴研究领域，目前在英国代表性不足。

项目成果

Structural dynamics in the evolution of SARS-CoV-2 spike glycoprotein

SARS-CoV-2 刺突糖蛋白进化的结构动力学

• DOI: http://dx.10.1038/s41467-023-36745-0
• 发表时间: 2023
• 期刊: Nature Communications
• 影响因子: 16.6
• 作者: Calvaresi V

Structural dynamics in the evolution of SARS-CoV-2 spike glycoprotein

SARS-CoV-2 刺突糖蛋白进化的结构动力学

• DOI: http://dx.10.21203/rs.3.rs-2049401/v1
• 发表时间: 2022
• 作者: Calvaresi V

Hydrogen/deuterium exchange-mass spectrometry of integral membrane proteins in native-like environments: current scenario and the way forward.

类天然环境中完整膜蛋白的氢/氘交换质谱：当前情况和前进方向。

• DOI: http://dx.10.1042/ebc20220173
• 发表时间: 2023
• 期刊: Essays in biochemistry
• 作者: Javed W

Chromatographic Phospholipid Trapping for Automated H/D Exchange Mass Spectrometry of Membrane Protein-Lipid Assemblies

用于膜蛋白-脂质组件自动 H/D 交换质谱分析的色谱磷脂捕获

• 发表时间: 2023
• 期刊: Analytical Chemistry
• 影响因子: 7.4
• 作者: Dietmar Hammerschmid

Integrating Hydrogen Deuterium Exchange-Mass Spectrometry with Molecular Simulations Enables Quantification of the Conformational Populations of the Sugar Transporter XylE

将氢氘交换质谱与分子模拟相结合，能够对糖转运蛋白 XylE 的构象群进行定量

• DOI: http://dx.10.1021/jacs.2c06148
• 发表时间: 2023
• 期刊: Journal of the American Chemical Society
• 影响因子: 15
• 作者: Jia R