Elucidating the mechanisms of protein secretion across the outer membrane by bacterial autotransporters

阐明细菌自转运蛋白跨外膜分泌蛋白质的机制

• 批准号: 10736193
• 负责人: James C. Gumbart
• 金额: $ 41.7万
• 依托单位: GEORGIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-18 至 2027-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10736193
• 关键词: Adhesions; Amino Acids; Anti-Bacterial Agents; Antibiotic Resistance; Atomic Force Microscopy; B-Lymphocytes; Bacterial Adhesins; Biological Assay; Bordetella pertussis; C-terminal; Cell surface; Cessation of life; Chemicals; Chicago; Classification; Collaborations; Complex; Coupling; Cryoelectron Microscopy; Data; Deuterium; Diffusion; Electrostatics; Environment; Escherichia coli; Exhibits; Extracellular Space; Free Energy; Goals; Grain; Gram-Negative Bacteria; Gram-Negative Bacterial Infections; Growth; Hybrids; Hydrogen; In Vitro; Individual; Infection; Infection Control; Intervention; Kinetics; Mass Spectrum Analysis; Mediating; Membrane; Methodology; Methods; Modeling; Molecular; N-terminal; Names; Pathogenicity; Pathway interactions; Peptide Hydrolases; Porosity; Positioning Attribute; Process; Protein Export Pathway; Protein Secretion; Proteins; Rationalization; Research; Resistance development; Side; Structure; Surface; System; Testing; Time; Tracheal Epithelium; Validation; Virulence; Virulence Factors; World Health Organization; Yersinia pestis; base; beta barrel; beta pleated sheet; crosslink; cytotoxic; deep learning; deep learning model; experimental study; extracellular; functional hypothalamic amenorrhea; in vivo; innovation; laboratory experiment; member; molecular dynamics; molecular scale; non-Native; novel; pathogen; pathogenic bacteria; periplasm; pertactin; prevent; priority pathogen; simulation; small molecule; tool

Project Summary/Abstract Pathogenic Gram-negative bacteria are collectively responsible for over 5 million deaths annually. A number of species also exhibit high levels of antibiotic resistance, comprising nine out of twelve members on the World Health Organization’s list of priority pathogens. Gram-negative bacterial infection is often mediated by so-called autotransporters, a class of proteins that cross the outer membrane to the extracellular space where they act as virulence factors, such as adhesins, proteases, and other harmful agents. Autotransporters consist of a translocator domain, which remains in the membrane, and a passenger domain, which secretes across the membrane to the other side, even without the use chemical energy, e.g., ATP. Targeting these autotransporters for inhibition represents a promising means of controlling infection while limiting the development of resistance. However, first, research into the molecular mechanisms of autotransporter folding, secretion, and expression beyond the cell surface is critically needed. This project will meet that need through three specific aims. In the first aim, how the passenger domain folds will be characterized, answering why folding in vivo is orders of magnitude faster than in vitro. The second aim focuses on the secretion of the passenger domain across the membrane through a hybrid-β-barrel of the translocator domain with BamA, the protein responsible for its membrane insertion. The pathway through the combined barrels will be determined, as well as the influence of the outer membrane on the process. In the third aim, another class of autotransporters, two-partner secretion systems, will be modeled, with a goal of identifying both commonalities and differences between them and other classes. The primary methodological tool to be used for this project is atomic-scale molecular dynamics (MD) simulations. These simulations will all be carried out in realistic environments, including an accurate model of the asymmetric Gram-negative outer membrane. Multiple innovative approaches will also be used, including deep learning for modeling the folding process, Markov state modeling for extracting kinetics information, and coarse- grained Brownian dynamics for observing spontaneous secretion. Close collaboration with multiple experimental labs will provide key inputs to and validation of the MD simulation results.

项目摘要/摘要 致病革兰氏阴性细菌每年统称超过500万人死亡。许多 物种还表现出较高的抗生素耐药性，在世界十二个成员中完成了九个 卫生组织的优先病原体清单。革兰氏阴性细菌感染通常是由所谓的 自动转运蛋白，一类蛋白质，它们越过外膜至细胞外空间 作为病毒因素，例如粘合剂，蛋白酶和其他有害药物。自动转移者由 保留在膜中的转运域和一个乘客域，该领域的秘密 即使没有使用化学能，例如ATP。针对这些自动转移者 对于抑制作用，代表了控制感染的承诺含义，同时限制了抗药性的发展。 但是，首先，研究自转运蛋白折叠，分泌和表达的分子机制 除了细胞表面之外，还需要非常需要。该项目将通过三个特定目标满足需求。在 第一个目的是如何对乘客域折叠进行表征，回答为什么在体内折叠为何折叠 比体外快的幅度。第二个目标重点是跨越乘客域的分泌 通过Bama的转运剂结构域的杂化型桶的膜，蛋白质负责 膜插入。将确定穿过组合桶的路径，以及 该过程的外膜。在第三个目标中，另一类自动转移者，两伴侣的分泌物 系统将被建模，其目标是确定它们与其他之间的共同点和差异 课程。该项目要使用的主要方法论工具是原子级分子动力学（MD） 模拟。这些模拟将全部在现实的环境中进行，包括准确的模型 非对称革兰氏阴性外膜。还将使用多种创新方法，包括 学习建模折叠过程，Markov状态建模用于提取动力学信息以及粗略 - 颗粒的布朗动力学用于观察赞助分泌。与多个实验的密切合作 实验室将为MD模拟结果提供关键输入和验证。

项目成果

From simple to complex: Reconstructing all-atom structures from coarse-grained models using cg2all.

从简单到复杂：使用 cg2all 从粗粒度模型重建全原子结构。

• DOI: 10.1016/j.str.2023.12.004
• 发表时间: 2024
• 期刊: Structure (London, England : 1993)
• 作者: Pang,YuiTik;Yang,Lixinhao;Gumbart,JamesC
• 通讯作者: Gumbart,JamesC