Bacterial Microcompartment Cargo Packing and Ultrastructure: High-Resolution Studies of Native Alpha-Carboxysomes

项目总结/摘要 细菌缺乏细胞器，但一些酶途径产生中间体，这些中间体对细胞器敏感。 损失或如果释放到胞质溶胶中可能对细胞有毒。大约20%的细菌包裹着潜在的 细菌微区室（BMC）内这些途径的危险或低效成分。BMC有 已被鉴定为许多种类，并发挥各种作用，从碳固定到小分子 新陈代谢.它们通常含有两种或两种以上的酶，可变数量的辅助和支架 蛋白质，以及在没有脂质帮助的情况下对小分子选择性渗透的致密蛋白质壳 膜的壳蛋白和壳的异质性组成在所有BMC中是保守的 到目前为止的所有功能。因为BMC可以让细菌在恶劣的环境中生存， 对人类健康、生态和传染病有着广泛的影响。BMCs也成为了 蛋白质工程，因为它们有可能包封替代药理学和 生物技术的应用，以及它们作为许多物种的重要组成部分的基本价值， 新陈代谢.尽管它们很重要，但结构的异质性阻碍了对它们的完全理解。 结构、超微结构和壳蛋白和货物的空间组织。研究 这里提出的目的是表征结构和组织的羧基体，一个模型BMC负责 用于蓝藻的碳固定。高分辨率冷冻电子断层扫描和子断层图像平均将 用于测定货物组织和壳的超微结构在体外和体内，保存和 表征结构的保守异质性。除了提供对BMC的新见解外， 生物学及其保守的复杂性，这项研究也将产生新的分析方法，可以应用 许多复杂的BMC和病毒系统。

Project Summary/Abstract Bacteria lack organelles, yet some enzymatic pathways generate intermediates which are either susceptible to loss or which could be toxic to the cell if released into the cytosol. Roughly 20% of bacteria encase the potentially dangerous or inefficient components of these pathways inside bacterial microcompartments (BMCs). BMCs have been characterized in a number of species, and serve a variety of roles from carbon fixation to small molecule metabolism. They typically contain two or more enzymes, a variable number of accessory and scaffolding proteins, and a compact protein shell that is selectively permeable to small molecules without the aid of a lipid membrane. The shell proteins and the heterogeneous composition of the shell are conserved across all BMCs of all functions characterized thus far. Because BMCs can allow bacteria to live in hostile environments, they have broad implications for human health, ecology and infectious disease. BMCs have also become a target of protein engineering due to their potential for enclosing cargos of choice for alternative pharmacological and biotechnological applications, as well as for their basic value as a critical component of many species’ metabolism. Despite their importance, structural heterogeneity has prevented a complete understanding of architecture, ultrastructure, and spatial organization of both the shell proteins and the cargo. The research proposed here seeks to characterize the structure and organization of carboxysomes, a model BMC responsible for carbon fixation in cyanobacteria. High-resolution cryo-electron tomography and sub-tomogram averaging will be used to determine cargo organization and shell ultrastructure in vitro and in vivo, preserving and characterizing the conserved heterogeneity of the structures. In addition to providing new insights into BMC biology and its conserved complexity, this research will also generate new analysis methods that can be applied to many complicated BMC and viral systems.