Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Molecules. 2019 Jan 30;24(3):496. doi: 10.3390/molecules24030496.
Advances in synchrotron technology are changing the landscape of macromolecular crystallography. The two recently opened beamlines at NSLS-II-AMX and FMX-deliver high-flux microfocus beams that open new possibilities for crystallographic data collection. They are equipped with state-of-the-art experimental stations and automation to allow data collection on previously intractable crystals. Optimized data collection strategies allow users to tailor crystal positioning to optimally distribute the X-ray dose over its volume. Vector data collection allows the user to define a linear trajectory along a well diffracting volume of the crystal and perform rotational data collection while moving along the vector. This is particularly well suited to long, thin crystals. We describe vector data collection of three proteins-Akt1, PI3Kα, and CDP-Chase-to demonstrate its application and utility. For smaller crystals, we describe two methods for multicrystal data collection in a single loop, either manually selecting multiple centers (using H108A-PHM as an example), or "raster-collect", a more automated approach for a larger number of crystals (using CDP-Chase as an example).
同步辐射技术的进步正在改变大分子晶体学的面貌。最近在 NSLS-II-AMX 和 FMX 开设的两条光束线提供了高通量微聚焦光束，为晶体学数据收集开辟了新的可能性。它们配备了最先进的实验站和自动化设备，可用于以前难以处理的晶体的数据收集。优化的数据收集策略允许用户根据需要将晶体定位，以将 X 射线剂量最佳地分布在晶体的体积上。向量数据收集允许用户沿着晶体具有良好衍射的体积定义线性轨迹，并在沿向量移动时执行旋转数据收集。这特别适用于长而细的晶体。我们描述了 Akt1、PI3Kα 和 CDP-Chase 三种蛋白质的向量数据收集，以展示其应用和实用性。对于较小的晶体，我们描述了两种在单个循环中收集多个晶体数据的方法，要么手动选择多个中心（以 H108A-PHM 为例），要么使用“光栅收集”，这是一种适用于更多晶体的更自动化的方法（以 CDP-Chase 为例）。
