Molecular architecture and mechanism of the chloroplast's beta-barrel assembly machinery (ChlAM)

叶绿体β-桶组装机器（ChlAM）的分子结构和机制

• 批准号: EP/Y036158/1
• 负责人: Jani Reddy Bolla
• 金额: $ 161.88万
• 依托单位: University of Oxford
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2024
• 资助国家: 英国
• 起止时间: 2024 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=EP%2FY036158%2F1
• 关键词: Molecular; architecture; mechanism; chloroplast; beta

A fundamental question in cell biology and biochemistry is how beta-barrel membrane proteins assemble into the outer membrane of Gram-negative bacteria and endosymbiotic organelles (mitochondria and chloroplasts). This is of profound importance because the function of many of these organelles depends upon the proper assembly of essential beta-barrel proteins involved in transporting ions, metabolites and proteins. Thus, a complete mechanistic understanding of this process may enable us to modify or control beta-barrel biogenesis to aid the development of specific therapies targeting resistant bacteria or mitochondrial diseases. Moreover, thisunderstanding will contribute to the development of crops with enhanced yields (since chloroplasts are responsible for photosynthesis), thereby addressing population needs and pressure from climate change.Our current knowledge of bacterial and mitochondrial beta-barrel assembly machineries makes it difficult to discern common underlying principles, generalities, and disparities among three model systems. Furthermore, many critical mechanistic steps remain unclear, including how the assembly machineries recognise, fold, and release substrates. This is primarily due to our inability to capture the transient interactions involved within this process. Based on my recent discovery that native mass spectrometry (MS) can efficiently capture these transient interactions and the information obtained that can guide structural analysis, I propose to decipher themolecular architecture and mechanism of the chloroplast beta-barrel assembly machinery and thereby contribute to a greater understanding of a unified mechanistic model. This ambitious project integrates several recent breakthrough discoveries in structural biology, such as native MS, top-down MS, mass photometry, and cryo-EM. Over the last year, I have laid the foundation and developed a pipeline to facilitate these studies and have demonstrated the project's overall feasibility.

细胞生物学和生物化学的一个基本问题是β-桶膜蛋白如何在革兰氏阴性细菌和内共生细胞器（线粒体和叶绿体）的外膜中组装成外膜。这是非常重要的，因为许多这些细胞器的功能取决于与运输离子，代谢物和蛋白质有关的必需β桶蛋白的适当组装。因此，对这一过程的完全机械理解可以使我们能够修改或控制β-桶生物发生，以帮助开发靶向抗性细菌或线粒体疾病的特定疗法。此外，这种理解将有助于产量增强的农作物的发展（因为叶绿体是造成光合作用的原因），从而满足人口需求和气候变化所带来的压力。我们当前对细菌和线粒体beta beta-barrel组装机械的了解使得很难辨别出常见的基本原理，一般性，三种模型和模型。此外，许多关键的机械步骤尚不清楚，包括装配机器如何识别，折叠和释放底物。这主要是由于我们无法捕获此过程中涉及的瞬态相互作用。基于我最近发现，天然质谱法（MS）可以有效地捕获这些瞬态相互作用以及可以指导结构分析的信息，我建议解解叶绿体β-桶式装配机械的分子结构和机制，从而有助于对统一机械模型的更多了解。这个雄心勃勃的项目整合了结构生物学中最近的几个突破性发现，例如天然MS，自上而下的MS，质量光度法和冷冻EM。在过去的一年中，我奠定了基础，并开发了一条管道来促进这些研究，并证明了该项目的整体可行性。