Structural studies of bacteriophage T4 and their potential medical applications

噬菌体T4的结构研究及其潜在的医学应用

• 批准号: 8959762
• 负责人: MICHAEL G ROSSMANN
• 金额: $ 65.26万
• 依托单位: PURDUE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-12-15 至 2020-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8959762
• 关键词: Acute; Anti-Bacterial Agents; Antibiotics; Antigens; Bacteria; Bacteriophage T4; Bacteriophages; Binding; Biochemical; Biological; Biological Models; Capsid; Capsid Proteins; Cells; Collaborations; Complex; Cryoelectron Microscopy; Crystallography; DNA Packaging; Data; Electron Microscopy; Electrons; Engineering; Epitopes; Eukaryotic Cell; Fiber; Foundations; Gene Delivery; Genome; Head; Infection; Isometric Exercise; Knowledge; Laboratories; Life Cycle Stages; Malignant Neoplasms; Medical; Microscopic; Modification; Molecular Biology; Motor; Mutation; Neck; Process; Prokaryotic Cells; Prolate; Protein Chemistry; Proteins; Recombinants; Research; Resistance; Resolution; Sampling; Staging; Structure; Surface; Tail; Techniques; Therapeutic; Toxin; Translating; Tube; Viral; Virus; cancer cell; cancer therapy; experience; gene therapy; genetic information; particle; pathogen; protein complex; protein protein interaction; public health relevance; reconstruction; tool; vaccine development

 DESCRIPTION (provided by applicant): Most bacterial viruses ("bacteriophages" or just "phages") are highly efficient in their ability to infect their often very specific bacterial hosts In general, only one or a few viral particles are necessary to successfully infect one bacterium. As a consequence there has been a long, unfulfilled hope that phages could be used as antibiotics. The need for alternative antibiotics is becoming ever more acute as common bacterial pathogens are developing resistant mutations, making it urgent to develop alternatives such as phage therapy. In addition, recombinant phages have been suggested as a means for gene therapy and as potential antigens for development of vaccines. Bacteriophage T4 has been a model system to study virus structure and function, having advanced biochemical principles of the assembly of biological complexes and protein-protein interactions. We have extensive experience in both structural and functional studies of phage T4. We now wish to expand this knowledge and make it available for potential medical applications. The plan is to analyze the structure and assembly of the capsid (Specific Aim 1), recognition of the host by the phage fibers (Specific Aim 2), the DNA packaging machine (Specific Aim 3), attachment of the packaging machine to the head (Specific Aim 4), and the mechanism of gene delivery (Specific Aim 5). These studies are directed towards engineering phage T4 for specific medical applications to target specific prokaryotic cells (as an antibiotic) and eukaryotic cells (such as cancer cells) for the delivery of toxins. In addition, we are engineering the surface of T4 to carr multiple copies of an epitope for vaccine development. Our primary tools will be molecular biology, protein chemistry, crystallography and electron microscopy. Molecular biology studies will produce pure samples in sufficient quantity for structural and functional studies. Crystallographic studies will be of protein components, providing three-dimensional information at near atomic resolution. Cryo-electron microscopic reconstructions will provide three-dimensional data on the organization of the protein components within the virus. Combining these techniques will generate "pseudo-atomic" resolution structures of the virus at different stages of its life cycle for functional interpretation and verification.

 描述（由申请人提供）：大多数细菌病毒（“噬菌体”或简称“噬菌体”）在感染其通常非常特异的细菌宿主的能力方面是高效的。通常，成功感染一种细菌仅需要一个或几个病毒颗粒。因此，长期以来一直存在着一个未实现的希望，即β-内酰胺酶可以用作抗生素。对替代抗生素的需求变得越来越迫切，因为常见的细菌病原体正在产生耐药性突变，迫切需要开发噬菌体疗法等替代药物。此外，已建议将重组人β-内酰胺酶作为基因治疗的手段和作为疫苗开发的潜在抗原。 噬菌体T4是研究病毒结构和功能的模型系统，具有生物复合物组装和蛋白质-蛋白质相互作用的先进生物化学原理。我们在噬菌体T4的结构和功能研究方面都有丰富的经验。我们现在希望扩展这些知识，并使其可用于潜在的医学应用。 该计划是分析衣壳的结构和组装（特异性目的1），噬菌体纤维对宿主的识别（特异性目的2），DNA包装机（特异性目的3），包装机与头部的连接（特异性目的4），以及基因递送机制（特异性目的5）。这些研究针对工程化噬菌体T4用于特定的医学应用，以靶向特定的原核细胞（作为抗生素）和真核细胞（如癌细胞）用于递送毒素。此外，我们正在对T4的表面进行工程改造，使其携带卡尔多个拷贝的表位用于疫苗开发。 我们的主要工具将是分子生物学，蛋白质化学，晶体学和电子显微镜。分子生物学研究将产生足够数量的纯样品用于结构和功能研究。晶体学研究将是蛋白质成分，提供近原子分辨率的三维信息。冷冻电子显微镜重建将提供病毒内蛋白质组分组织的三维数据。结合这些技术将产生病毒在其生命周期的不同阶段的“伪原子”分辨率结构，用于功能解释和验证。