Deciphering the molecular principles of bacterial metabolosome biogenesis

破译细菌代谢体生物发生的分子原理

• 批准号: BB/V009729/1
• 负责人: Luning Liu
• 金额: $ 87.75万
• 依托单位: University of Liverpool
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2021
• 资助国家: 英国
• 起止时间: 2021 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FV009729%2F1
• 关键词: Deciphering; molecular; principles; bacterial; metabolosome

Pathogenic bacteria, such as Salmonella, thrive in mammalian intestines and can cause severe health issues in human, including food poisoning, massive gut inflammation and cardiovascular disease. There were estimated 535,000 cases of Salmonella gastrointestinal infections worldwide in 2017, and 91,857 cases in the EU in 2018. Salmonella cells produce a specialised nano-scale organelle, known as the bacterial microcompartment. These organelles provide a suite of unique metabolic advantages that allow Salmonella to become the predominant species in the hostile environment of the host gut. The organelle uses a shell that is made of thousands of proteins to sequester multiple enzymes used for 1,2-propanediol utilisation (Pdu). This unique structure allows the Pdu organelles to protect bacterial cells from toxic metabolites and to enhance the cell's metabolism. Although the importance of Pdu organelles for the metabolism of bacterial pathogens is appreciated, little is known about how bacterial cells generate and then modulate these organelles to confer adaptive cellular metabolism to survive in the sophisticated gut environment.We have recently reported the exact protein stoichiometry of Pdu organelles and have established a new structural model of the organelle. We have also developed systems for tagging Pdu proteins with fluorescent markers and depleting target proteins, so that we can track specific building proteins using microscopes and study their functions in bacterial cells. Using the developed systems, we have discovered that the cargo enzymes and shell proteins self-assemble independently in Salmonella. We have also shown that the locations and movement of Pdu organelles are confined within the bacterial cell. Standing on these exciting scientific and technical breakthroughs and an established research team with complementary expertise, we now aim to do an in-depth exploration of how Pdu organelles are synthesized and how the organisation of Pdu organelles is coordinated within the Salmonella cell. We will first determine the multi-step assembly that individual building proteins undergo to form higher-ordered assemblies, identify the proteins that make up the enzyme and shell assemblies, and elucidate how enzyme and shell assemblies associate together to form an intact organelle. In the second section of our programme, we will characterise the structures and functions of small linker proteins that drive the assembly of cargos to have a "liquid-like" dynamic phase and ascertain that the phase separation mechanism is vital for mediating the protein interactions and formation of a functional protein organelle. Finally, we will use state-of-the-art fluorescence microscopy to probe how the Pdu organelles are generated, located, and modulated to perform such important functions in bacterial cells.This ambitious and multidisciplinary research project has both fundamental and applied significance. Pdu MCPs represents an ideal model system for uncovering the principles of protein self-assembly and the generation of multi-protein complexes in biology. We will learn the basic physics and chemistry of how thousands of proteins assemble together to build a functional entity within a bacterial cell, and determine how the cell precisely and efficiently controls the formation and function of metabolic organelles. We anticipate that our findings will provide a deeper understanding of the biosynthesis and maintenance of natural bacterial organelles and protein assemblies. The research may inform strategies for the engineering of biological "factories" for the enhancement of cell metabolism and energy production in diverse biotechnological applications. Moreover, the essential protein-protein interactions that we find are required to mediate the assembly of Pdu organelles could represent novel therapeutic targets to disrupt the production of Pdu organelles and thus ablate the ability of Salmonella to thrive in the human gut.

致病细菌（例如沙门氏菌）在哺乳动物肠道中繁衍生息，并可能在人类中引起严重的健康问题，包括食物中毒，大规模的肠道炎症和心血管疾病。 2017年，全球估计有535,000例沙门氏菌胃肠道感染，2018年欧盟有91,857例。沙门氏菌细胞产生了一种专业的纳米尺度细胞器，称为细菌微型室。这些细胞器提供了一系列独特的代谢优势，使沙门氏菌能够成为宿主肠敌对环境中主要物种。细胞器使用由数千种蛋白质制成的壳来隔离用于1,2-丙二醇利用率（PDU）的多种酶。这种独特的结构使PDU细胞器可以保护细菌细胞免受有毒代谢物的影响，并增强细胞的代谢。尽管人们对PDU细胞器对细菌病原体的代谢的重要性得到了赞赏，但对细菌细胞如何产生，然后调节这些细胞器来赋予适应性细胞代谢以生存以在精致的肠道环境中生存。我们最近报告了PDU Organelles的确切蛋白质量表，并建立了一个新的结构模型。我们还开发了用于用荧光标记和耗尽靶蛋白标记PDU蛋白的系统，以便我们可以使用显微镜跟踪特定的建筑蛋白质并研究其在细菌细胞中的功能。使用开发的系统，我们发现货物酶和壳蛋白在沙门氏菌中独立地自组装。我们还表明，PDU细胞器的位置和运动局限于细菌细胞内。站在这些令人兴奋的科学和技术突破以及具有互补专业知识的知名研究团队的基础上，我们现在旨在深入探索PDU细胞器如何合成，以及在沙门氏菌细胞中如何协调PDU细胞器的组织。我们将首先确定多步组件，即单个建筑蛋白会经历以形成高阶组件，识别构成酶和壳体组件的蛋白质，并阐明酶和壳体组件如何将酶和外壳组合在一起形成完整的细胞器。在我们程序的第二部分中，我们将表征小连接蛋白的结构和功能，这些连接蛋白驱动千片的组装具有“液体样”动态相，并确定相位分离机制对于介导蛋白质相互作用和功能性蛋白质细胞器的形成至关重要。最后，我们将使用最先进的荧光显微镜来探测如何在细菌细胞中产生，定位和调节PDU细胞器。这些雄心勃勃的多学科研究项目具有基本性和具有重要意义。 PDU MCPS代表了一个理想的模型系统，用于揭示蛋白质自组装原理和生物学中多蛋白质复合物的产生。我们将学习成千上万种蛋白质聚集在一起以在细菌细胞中构建功能实体的基本物理和化学，并确定细胞如何精确有效地控制代谢细胞器的形成和功能。我们预计我们的发现将为天然细菌细胞器和蛋白质组件的生物合成和维持提供更深入的了解。这项研究可能为生物“工厂”工程的策略提供依据，以增强各种生物技术应用中细胞代谢和能源生产。此外，我们发现的必需蛋白质 - 蛋白质相互作用是介导PDU细胞器组装的必需蛋白质蛋白质相互作用，可以代表新的治疗靶标，以破坏PDU细胞器的产生，从而消除沙门氏菌在人肠道中繁殖的能力。

项目成果

Probing the Internal pH and Permeability of a Carboxysome Shell.

• DOI: 10.1021/acs.biomac.2c00781
• 发表时间: 2022-10-10
• 期刊: BIOMACROMOLECULES
• 影响因子: 6.2
• 作者: Huang, Jiafeng;Jiang, Qiuyao;Yang, Mengru;Dykes, Gregory F.;Weetman, Samantha L.;Xin, Wei;He, Hai-Lun;Liu, Lu-Ning
• 通讯作者: Liu, Lu-Ning

Cryo-EM structure of a monomeric RC-LH1-PufX supercomplex with high-carotenoid content from Rhodobacter capsulatus

• DOI: 10.1016/j.str.2023.01.006
• 发表时间: 2023-03-02
• 期刊: STRUCTURE
• 影响因子: 5.7
• 作者: Bracun, Laura;Yamagata, Atsushi;Liu, Lu-Ning
• 通讯作者: Liu, Lu-Ning

Producing fast and active Rubisco in tobacco to enhance photosynthesis.

• DOI: 10.1093/plcell/koac348
• 发表时间: 2023-02-20
• 期刊: PLANT CELL
• 影响因子: 11.6
• 作者: Chen, Taiyu;Riaz, Saba;Davey, Philip;Zhao, Ziyu;Sun, Yaqi;Dykes, Gregory F.;Zhou, Fei;Hartwell, James;Lawson, Tracy;Nixon, Peter J.;Lin, Yongjun;Liu, Lu-Ning
• 通讯作者: Liu, Lu-Ning

Chemoautotrophic production of gaseous hydrocarbons, bioplastics and osmolytes by a novel Halomonas species.

• DOI: 10.1186/s13068-023-02404-1
• 发表时间: 2023-10-11
• 期刊: Biotechnology for biofuels and bioproducts

Incorporation of Functional Rubisco Activases into Engineered Carboxysomes to Enhance Carbon Fixation.

• DOI: 10.1021/acssynbio.1c00311
• 发表时间: 2022-01-21
• 期刊: ACS synthetic biology
• 影响因子: 4.7
• 作者: Chen T;Fang Y;Jiang Q;Dykes GF;Lin Y;Price GD;Long BM;Liu LN
• 通讯作者: Liu LN