MICROFOCUS X-RAY DATA COLLECTION WITH DIFFRACTION-CAPABLE MICROFLUIDIC CHIPS

使用具有衍射功能的微流控芯片进行微焦 X 射线数据收集

• 批准号: 7955162
• 负责人: JAMES BERGER
• 金额: $ 1.93万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2010-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7955162
• 关键词: Biological; Complex; Computer Retrieval of Information on Scientific Projects Database; Crystallization; DNA; DNA biosynthesis; Data Collection; Funding; Grant; In Situ; Institution; Membrane Proteins; Microfluidics; Molecular Conformation; Research; Research Personnel; Resources; Roentgen Rays; Sampling; Source; Structure; System; United States National Institutes of Health; Virulence Factors; beamline; cryogenics; design; interest; macromolecular assembly; molecular assembly/self assembly; research study; structural biology

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Microfocus X-ray Data Collection with Diffraction-Capable Microfluidic Chips and Microcrystals of Large Molecular Assemblies We have recently developed microfluidic crystallization chips designed for in-situ diffraction data collection. These chips have been designed for low-volume crystallization experiments (10nl sample per sample chamber) and to minimize x-ray background scatter from the chip material. Sections can be removed from chips, cryoprotectants can be added, and cryogenic data collection is possible. Crystals grown in these chips are typically small (40x40x40 ¿m) and data collection from them will benefit from the use of microfocus X-ray sources to further reduce background X-ray scatter. This subproject aims to demonstrate the utility of the NE-CAT microfocus X-ray beamline for data collection from microfluidic chips and determine structures of samples of broad biological interest including DNA replication complexes, membrane proteins and bacterial virulence factors. We also have been studying large macromolecular assemblies that remodel and replicate DNA. Many of these systems assume multiple conformational states during their catalytic cycle, making them challenging structural targets, and prone to forming microcrystals despite extensive optimization efforts.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 使用具有衍射功能的微流控芯片和大分子组件微晶进行微焦点 X 射线数据采集 我们最近开发了专为原位衍射数据收集而设计的微流体结晶芯片。这些芯片专为小体积结晶实验（每个样品室 10nl 样品）而设计，可最大程度地减少芯片材料的 X 射线背景散射。可以从芯片上移除切片，可以添加冷冻保护剂，并且可以收集低温数据。这些芯片中生长的晶体通常很小（40x40x40 µm），并且从它们收集的数据将受益于微焦点 X 射线源的使用，以进一步减少背景 X 射线散射。该子项目旨在展示 NE-CAT 微焦点 X 射线光束线在微流控芯片数据收集中的实用性，并确定具有广泛生物学意义的样品结构，包括 DNA 复制复合物、膜蛋白和细菌毒力因子。 我们还一直在研究重塑和复制 DNA 的大分子组装体。 许多这些系统在催化循环过程中呈现多种构象状态，这使得它们具有挑战性的结构目标，并且尽管进行了大量的优化工作，仍然容易形成微晶。