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

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 可衍射型微流控芯片和大分子组装微晶的微焦点X射线数据采集 我们最近开发了用于原位衍射数据采集的微流控结晶芯片。这些芯片设计用于低容量结晶实验(每个样品室10nl样品)，并将来自芯片材料的X射线背景散射降至最低。可以从芯片上移除切片，可以添加低温保护剂，还可以收集低温数据。在这些芯片中生长的晶体通常很小(40x40x40？m)，从这些芯片中收集数据将受益于使用微焦点X射线源来进一步减少背景X射线散射。这一分项目旨在展示NE-CAT微焦点X射线光束线用于从微流控芯片收集数据的用途，并确定具有广泛生物学意义的样品的结构，包括DNA复制复合体、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。其中许多体系在催化循环中呈现多种构象状态，使它们具有挑战性的结构目标，尽管进行了广泛的优化工作，但仍容易形成微晶。

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.