Structure, Function, and Disruption of Microbial Amyloid Assembly and Biofilm For

微生物淀粉样蛋白组装和生物膜的结构、功能和破坏

基本信息

批准号：
7981064
负责人：
Lynette S Cegelski
金额：
$ 237万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-30 至 2015-06-30
项目状态：
已结题

项目摘要

DESCRIPTION (Provided by the applicant) Abstract: Structure, Function, and Disruption of Microbial Amyloid Assembly and Biofilm Formation Biofilm bacteria are wreaking havoc in the clinic. Our current arsenal of antibiotics is comprised of variations on the single theme of cell killing, i.e. targeting processes essential to bacterial viability, yet biofilms exhibit reduced sensitivity to conventional antibiotics and host defenses, and have emerged as virulence hallmarks of serious and persistent infectious diseases, including cystic fibrosis, urinary tract infection, endocarditis, and catheter infections. Improved biofilm models are crucial to understanding function and driving the design of new anti-infectives. In this New Innovator Award application I present an unconventional research plan designed to enable and deliver breakthrough discoveries needed to transform biofilm descriptors from vague terms like "glue" and "slime" to quantitative descriptions based on chemical composition and molecular architecture. I achieve this by integrating new whole-cell solid-state NMR strategies with chemical biology, microscopy, and biochemical techniques. From a "systems perspective," carbon and nitrogen NMR spectra of intact biofilms will permit a sum-of-all-parts analysis, while selective recoupling measurements, together with steady-state and transient biosynthetic labeling, will assign unique molecular signatures to permit biofilm profiling as a function of time and organism. Bacterial amyloids are prevalent in biofilms among diverse phyla and contribute to biofilm formation and virulence by uropathogenic E. coli. We will dissect in atomic-level detail the molecular basis of amyloid biogenesis. We will then establish how bacteria use these structures as building blocks in the construction of biofilm architectures. We will also be engaged in identifying compounds that interfere with assembly processes and will examine the nature of disruption to improve our understanding of the multi-protein amyloid machinery and biofilm manufacturing in E. coli vis-¿-vis chemical genetics. In a broader context, results of this project over the next five years will establish an experimental basis for quantitatively characterizing heterogeneous, insoluble, noncrystalline assemblies that contribute to human infectious diseases. Public Health Relevance: This era may come to be remembered as one in which infectious diseases made a dramatic worldwide resurgence, owing to the challenge of treating persistent biofilmassociated infectious diseases, the increasing rise of antibiotic resistance, and the dwindling number of candidate antibiotics in the drug development pipeline. We propose an unconventional and innovative approach that will enable transformative discoveries to understand at a molecular and atomic level how bacteria assemble complex heteropolymeric extracellular structures, including functional amyloid fibers, and how bacteria use these building blocks to construct organized biofilm architectures. We will also be engaged in identifying small molecules to interfere with assembly processes to drive the development of new anti-amyloid, anti-biofilm, and anti-virulence therapeutics.

描述（申请人提供）摘要：微生物淀粉样蛋白组装和生物膜形成生物膜细菌的结构，功能和破坏在诊所造成了严重破坏。我们目前的抗生素武器包括对细胞杀戮的单一主题的变化，即针对细菌生存能力所必需的靶向过程，但生物膜暴露了对常规抗生素和宿主防御剂的敏感性降低，并且已成为严重和持续性感染疾病的病毒标志，包括细胞和静脉内感染，包括囊肿性感染，尿布式感染，尿布型，尿布型，尿布型，尿布型，尿布型，尿布式，尿液尿布型。改进的生物膜模型对于理解功能和推动新反感染物的设计至关重要。在这个新的创新奖应用程序中，我介绍了一项非常规研究计划，旨在使并提供突破性发现，从而将生物膜描述从“胶水”和“粘液”等投票术语转变为基于化学组成和分子体系结构的定量描述。我通过将新的全细胞固态NMR策略与化学生物学，显微镜和生化技术相结合来实现这一目标。从“系统的角度”来看，完整生物膜的碳和氮NMR光谱将允许全方位的分析，而选择性的重新耦合测量以及稳态和瞬态生物合成标记，将分配独特的分子特征，以使生物膜分析能够作为时间和组织的功能进行生物膜分析。细菌淀粉样蛋白在潜水员的生物膜中普遍存在，并通过肝癌大肠杆菌有助于生物膜形成和病毒。我们将在原子级的细节中剖析淀粉样生物发生的分子基础。然后，我们将建立细菌如何将这些结构用作生物膜体系结构的建设中的基础。我们还将参与识别干扰组装过程的化合物，并将检查中断的性质，以提高我们对大肠杆菌化学遗传学的多蛋白质淀粉样蛋白机械和生物膜制造的理解。在更广泛的背景下，未来五年的该项目的结果将建立一个实验基础，以定量表征有助于人类感染疾病的异质，不溶性，非晶体组件。公共卫生相关性：由于治疗持续存在的生物膜相关的感染性疾病的挑战，抗生素抗性的增加以及药物开发中的候选抗生素数量的越来越多，这种擦除可能会被人们铭记为一种传染病在全球范围内戏剧性的复发。我们提出了一种非常规和创新的方法，该方法将使变革性发现能够在分子和原子级别理解细菌如何组装复合的复合杂质外结构，包括功能性淀粉样蛋白纤维，以及细菌如何使用这些基础来构建有组织的生物纤维结构。我们还将参与鉴定小分子以干扰组装过程，以推动新的抗淀粉样蛋白，抗生物膜和抗病毒疗法的发展。