Probing the architecture, assembly, and function of amyloid-polysaccharide entanglements in bacterial biofilms

探究细菌生物膜中淀粉样蛋白-多糖缠结的结构、组装和功能

• 批准号: 10605820
• 负责人: Schuyler A. Chambers
• 金额: $ 6.91万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-01 至 2025-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10605820
• 关键词: Acceleration; Address; Adhesives; Ammonium; Amyloid; Amyloid Proteins; Amyloid fibers; Anti-Bacterial Agents; Anti-Infective Agents; Antibiotics; Architecture; Arginine; Bacteria; Bacterial Polysaccharides; Binding; Biochemical; Biological Assay; Biophysics; Biopolymers; Cell surface; Cells; Cellulose; Chemicals; Communities; Creativeness; Detection; Development; Diffusion; Disease; Disinfectants; Escherichia coli; Evaluation; Event; Exhibits; Extracellular Matrix; Extracellular Protein; Fellowship; Fluorescence Microscopy; Foundations; Future; Human Microbiome; Infection; Institution; Investigation; Isotope Labeling; Link; Magnetic Resonance; Mechanics; Mentorship; Microbial Biofilms; Modification; Molecular; NMR Spectroscopy; Nature; Nuclear Magnetic Resonance; Organism; Pathogenesis; Pathology; Polymers; Polysaccharides; Production; Research; Research Project Grants; Role; Salmonella; Salmonella enterica; Sampling; Structure; Symbiosis; Testing; Time; Tissues; Training; Universities; Urinary tract; Vancomycin; Visualization; Western Blotting; Work; bacterial community; cell community; clinically relevant; cohesion; commensal bacteria; design; efficacy evaluation; exhaust; experimental study; extracellular; immunoregulation; in vivo; member; microbial; microbiome; multidisciplinary; mutant; novel; pathogen; pathogenic bacteria; phosphoethanolamine; pressure; recruit; solid state nuclear magnetic resonance

Project Summary Bacteria are most commonly found in nature in multicellular communities termed biofilms. Biofilms are formed when bacteria synthesize, secrete, and enmesh themselves with diverse biopolymers. Beneficial bacteria in the microbiome assemble biofilms, while biofilms are unfortunately also linked to difficult-to-treat infections that exhibit increased tolerance to antibacterials and can exhaust treatment options. However, there are no blueprints for how bacteria build these tissue-like architectures and uncovering these details can accelerate discovery of new anti-infectives. E. coli, in particular, are normal residents in the healthy microbiome, but emerge as pathogens when they egress and colonize the urinary tract. E. coli, Salmonella species and other Gram-negative organisms harness specific amyloid and polysaccharide machinery to elaborate mechanically robust extracellular matrix architectures resembling baskets and blankets that surround cells and drive the formation of tissue-like biofilms. Due to the complexity of biopolymer composites, there are significant challenges associated with studying their structure and function, yet the ubiquity of these biopolymers makes them of high importance for study. This research plan is directed to test molecular hypotheses for how bacteria employ curli and phosphoethanolamine cellulose, a newly discovered chemically modified form of cellulose, to enmesh themselves in extracellular matrix (ECM). The research plan will test hypotheses regarding functional roles that we propose are ascribed to the zwitterionic phosphoethanolamine modification. Aim 1 is directed to evaluate the temporal and spatial developments of matrix assembly beyond the bacterial cell surface using fluorescence microscopy and creative functional biochemical assays. Aim 2 will implement a strategically designed solid-state nuclear magnetic resonance (NMR) approach to detect molecular contacts between polysaccharides and protein amyloids that are responsible for matrix cohesion. The functional benefit of ECM biopolymers will be determined in Aim 3, where clinically relevant antibiotics and a novel vancomycin-conjugate will be evaluated for efficacy against pEtN cellulose and curli containing biofilms. This work promises to formulate a molecular foundation for future avenues of inquiry at the host-pathogen interface, involving possible immunomodulatory roles of bacterial polysaccharides and amyloids, and possible biopolymer contributions to microbiome symbiosis and amyloid- associated disease pathologies. The fellowship candidate will receive significant training in solid-state NMR spectroscopy to study molecular interactions within E. coli biofilms and biochemical approaches to investigate bacterial communities. The considerable support and mentorship structure provided through this fellowship, the research sponsor (Prof. Lynette Cegelski) and institution (Stanford University) will facilitate the professional development of the fellowship applicant and the rigorous scientific investigation of the proposed research.

项目摘要 细菌在自然界中最常见于称为生物膜的多细胞群落中。形成生物膜 当细菌合成，分泌，并与各种生物聚合物纠缠在一起时。有益菌 微生物组组装生物膜，而不幸的是，生物膜也与难以治疗的感染有关， 表现出对抗菌药物的耐受性增加，并可能耗尽治疗选择。然而，没有蓝图 了解细菌是如何构建这些组织样结构的，揭示这些细节可以加速发现 新的抗感染药E.特别是大肠杆菌，它们是健康微生物组中的正常居民， 当病原体外出并定植在泌尿道时。E.大肠杆菌、沙门氏菌和其他革兰氏阴性菌 生物体利用特定的淀粉样蛋白和多糖机制， 细胞外基质结构类似于篮子和毯子，包围细胞并驱动细胞的形成。 组织样生物膜。由于生物聚合物复合材料的复杂性，存在相关的重大挑战 研究它们的结构和功能，但这些生物聚合物的普遍存在使它们具有高度的重要性 为了学习这项研究计划旨在测试细菌如何利用curli的分子假设， 磷酸乙醇胺纤维素，一种新发现的纤维素的化学改性形式， 细胞外基质（ECM）。该研究计划将测试有关职能作用的假设， 我们建议归因于两性离子磷酸乙醇胺改性。目标1是评估 使用荧光的细菌细胞表面外基质组装的时间和空间发展 显微镜和创造性的功能生化分析。Aim 2将实现一个战略性设计的固态 核磁共振（NMR）方法检测多糖和蛋白质之间的分子接触 淀粉样蛋白负责基质的凝聚。将确定ECM生物聚合物的功能益处 在目标3中，将评价临床相关抗生素和新型万古霉素缀合物的疗效 针对含有pEtN纤维素和卷曲的生物膜。这项工作有望制定一个分子基础， 宿主-病原体界面的未来研究途径，包括细菌的可能免疫调节作用 多糖和淀粉样蛋白，以及可能的生物聚合物对微生物组共生和淀粉样蛋白的贡献- 相关疾病病理学。研究金候选人将接受固体核磁共振方面的重要培训 光谱研究分子间的相互作用E.大肠杆菌生物膜和生化方法来研究 细菌群落通过该研究金提供的大量支持和指导结构， 研究赞助商（Lynette Cegelski教授）和机构（斯坦福大学）将促进专业人士 研究金申请人的发展和对拟议研究的严格科学调查。