Structure, Dynamics and Activity of the Bacterial Secretosome

细菌分泌体的结构、动力学和活性

• 批准号: BB/Y004981/1
• 负责人: Ian Collinson
• 金额: $ 72.51万
• 依托单位: University of Bristol
• 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=BB%2FY004981%2F1
• 关键词: Structure; Dynamics; Activity; Bacterial; Secretosome

All cells are surrounded by membranes, made up from a double layer of fatty molecules called phospholipids. Cell membranes act as a molecular "skin", keeping the cell's insides in, and separating different biochemical reactions. This barrier needs to be traversed in a controlled manner to allow the import of nutrients, and removal of waste products, and for communication with the outside world. This is achieved by a wide range of proteins that reside in the various membranes. We understand a great deal about the diverse biological functions that membrane proteins bestow, such as transport, respiration, photosynthesis. However, we know very little about how membranes are formed, or about the necessary transport of proteins across or into membranes during their biogenesis.Our proposal aims to gain a complete understanding about how proteins are exported-protein secretion. The secretory ('Sec' for short) machinery is essential for life-for every cell in every organism. The project concerns this process, in bacterial cells. Bacteria secrete proteins for a wide range of membrane and extracellular activities including for: cell adherence, pathogenicity, the degradation of antibiotics, including also the biogenesis of the protective cell wall or envelope. A major class of bacteria known as Gram-negatives, possess a cell wall composed of a periplasm with a peptidoglycan (PG) layer, surrounded by an outer-membrane. The biogenesis of the cell wall is dependent on protein secretion from the cell interior through the Sec machinery. Proteins of the periplasm can readily fold and remain there, while those destined for the cell surface are delivered to another transport machine called the BAM complex, for incorporation into the outer-membrane. The journey from the inner- to the outer-membrane through the envelope, is poorly understood. We are concerned with the question: how do proteins make their way rapidly and efficiently through this very crowded and challenging environment? We have discovered that the protein transport machineries of the inner- (Sec) and outer-membranes (BAM) as well as various accessory factors concerned with quality control interact to form an assembly that spans the bacterial envelope, which we have called the bacterial secretosome. The existence of a contiguous conduit through the envelope will have far reaching implications for our understanding of outer-membrane biogenesis, including an answer to the question posed above.The project will harness complementary expertise in biochemistry and new breakthrough technologies for high resolution imaging (electron microscopy) and accurate mass measurements (mass spectrometry). This powerful combination will allow us to examine the activity, structure and dynamics of the assembly in order to understand how it works-how proteins are delivered through the inner-membrane to the periplasm and the outer-membrane. This knowledge is important because it will enlighten our understanding of a fundamental aspect of bacterial physiology, and inter-membrane transport throughout biology. Moreover, new advances in our understanding of the bacterial secretosome will help develop strategies to compromise envelope biogenesis and its regenerative capabilities-essential for survival. This would generate much needed ammunition in our fight against antimicrobial resistance (AMR).

所有的细胞都被膜包围，膜由一层叫做磷脂的脂肪分子组成。细胞膜扮演着分子“皮肤”的角色，将细胞内部隔离开来，分离不同的生化反应。这一障碍需要以有控制的方式跨越，以允许营养物质的进口、废物的清除以及与外部世界的沟通。这是通过驻留在各种膜中的各种蛋白质来实现的。我们对膜蛋白所具有的运输、呼吸、光合作用等多种生物学功能有了很深的了解。然而，我们对膜是如何形成的，或者关于蛋白质在生物发生过程中必要的跨膜或进入膜的运输知之甚少。我们的建议旨在全面了解蛋白质是如何输出的--蛋白质分泌。分泌(简称SEC)机制对生命至关重要--对每个有机体中的每一个细胞都是如此。该项目涉及细菌细胞中的这一过程。细菌为广泛的细胞膜和细胞外活动分泌蛋白质，包括：细胞黏附、致病性、抗生素的降解，还包括保护细胞壁或包膜的生物发生。革兰氏阴性杆菌是一类主要的细菌，它的细胞壁由外膜包围的周质和肽聚糖(PG)层组成。细胞壁的生物发生依赖于细胞内部通过SEC机制分泌的蛋白质。周质的蛋白质可以很容易地折叠并留在那里，而那些被送到细胞表面的蛋白质则被送到另一台称为BAM复合体的运输机器上，以便结合到外膜中。通过包膜从内膜到外膜的旅程，人们知之甚少。我们关心的问题是：蛋白质如何快速有效地通过这个非常拥挤和具有挑战性的环境？我们发现，内膜(SEC)和外膜(BAM)的蛋白质运输机制以及与质量控制有关的各种辅助因素相互作用，形成一个跨越细菌被膜的组装，我们称之为细菌分泌体。通过包膜的连续管道的存在将对我们对外膜生物发生的理解产生深远的影响，包括对上述问题的回答。该项目将利用生物化学方面的互补专业知识和高分辨率成像(电子显微镜)和精确质量测量(质谱学)的新突破技术。这种强大的结合将使我们能够检查组装的活性、结构和动力学，以了解它是如何工作的--蛋白质是如何通过内膜输送到周质和外膜的。这一知识很重要，因为它将启发我们对细菌生理学的一个基本方面的理解，以及整个生物学中的膜间运输。此外，我们对细菌分泌体理解的新进展将有助于制定策略，以损害包膜生物发生及其再生能力--这是生存所必需的。这将在我们对抗抗菌素耐药性(AMR)的斗争中产生急需的弹药。