'Structure and function of the R-body, a piston-like nanomachine

“活塞式纳米机器 R 体的结构和功能

• 批准号: 10166870
• 负责人: Justin M Kollman
• 金额: $ 33.04万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-06-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10166870
• 关键词: 3-Dimensional; Architecture; Bacteria; Behavior; Biological; Biological Process; Biosensor; Biotechnology; Caliber; Cells; Chemicals; Contracts; Cryo-electron tomography; Cryoelectron Microscopy; Devices; Disulfides; Generations; Histidine; Hybrids; Image; Individual; Kinetics; Lasers; Maps; Measurement; Measures; Membrane; Microscopy; Modeling; Molecular; Molecular Conformation; Mutagenesis; Mutation; Nature; Optics; Paramecium; Polymers; Property; Protein Subunits; Proteins; Puncture procedure; Recombinants; Resolution; Rod; Rupture; Site; Structural Models; Structure; System; Testing; Therapeutic; Variant; Work; X-Ray Crystallography; base; biophysical properties; conformational conversion; crosslink; mutant; nanomachine; novel; optical traps; particle; solid state nuclear magnetic resonance; submicron; three dimensional structure; weapons

Project Summary R-bodies are self-assembling protein polymers from bacterial ensymbionts of 'killer' Paramecium strains that confer to the host the ability to kill competing strains. The tightly coiled sub-micron sized R-body undergoes a massive pH induced conformational change into elongated rods tens of microns long, generating forces sufficient to rupture biological membranes. Purified recombinant R-bodies can undergo many successive rounds of piston-like extension and contraction using only the chemical energy from pH change. These robust nanomachines have enormous potential for biotechnological and therapeutic applications. Their ability to behave as sensitive switches and to do work on biological structures can be adapted for many functions - they have, for example, been proposed as systems for phagosomal delivery of bioactive molecules in a way that would mimic their biological function. The ability to tune R-body behavior was recently demonstrated by a panel of point mutants that alter the triggering pH for extension. However, the lack of detailed molecular understanding of their assembly and the mechanism of the pH-sensitive piston-like extension are major roadblocks to developing R-bodies for novel applications. This proposal aims to determine the molecular architecture of R-bodies using a hybrid structural approach, and to measure their biophysical properties using optical trapping force measurements. Visualizing the three-dimensional structure of R-bodies in coiled and extended states will allow us to define the molecular mechanism of their piston-like behavior, laying the groundwork for tuning the structures to new functions.

项目概要 R-体是来自“杀手”草履虫菌株的细菌共生体的自组装蛋白质聚合物， 赋予宿主杀死竞争菌株的能力。紧密卷绕的亚微米尺寸的 R 体经历了 大量 pH 诱导构象变化为数十微米长的细长杆，产生力 足以使生物膜破裂。纯化的重组 R 体可以经历许多连续的过程 仅利用 pH 值变化产生的化学能进行多轮活塞式伸展和收缩。这些坚固的 纳米机器在生物技术和治疗应用方面具有巨大的潜力。他们的能力 充当敏感开关并在生物结构上进行工作可以适应许多功能 - 它们 例如，已被提议作为生物活性分子的吞噬体递送系统，其方式为 会模仿它们的生物学功能。最近通过一项研究证明了调节 R 体行为的能力 一组改变延伸触发 pH 值的点突变体。但缺乏详细的分子 了解它们的组装和 pH 敏感的活塞状延伸的机制是重要的 开发新应用 R 体的障碍。该提案旨在确定分子 使用混合结构方法构建 R 体结构，并使用以下方法测量其生物物理特性 光学捕获力测量。可视化卷曲和 R 体的三维结构 扩展态将使我们能够定义其类似活塞行为的分子机制，奠定了 为调整结构以适应新功能奠定基础。