Hutin Stephanie, Santoni Gianluca, Zander Ulrich, Foos Nicolas, Aumonier Sylvain, Gotthard Guillaume, Royant Antoine, Mueller-Dieckmann Christoph, Leonard Gordon
European Synchrotron Radiation Facility, Structural Biology Group;
European Synchrotron Radiation Facility, Structural Biology Group.
J Vis Exp. 2019 Mar 19(145). doi: 10.3791/58594.
X-ray crystallography is the major technique used to obtain high resolution information concerning the 3-dimensional structures of biological macromolecules. Until recently, a major requirement has been the availability of relatively large, well diffracting crystals, which are often challenging to obtain. However, the advent of serial crystallography and a renaissance in multi-crystal data collection methods has meant that the availability of large crystals need no longer be a limiting factor. Here, we illustrate the use of the automated MeshAndCollect protocol, which first identifies the positions of many small crystals mounted on the same sample holder and then directs the collection from the crystals of a series of partial diffraction data sets for subsequent merging and use in structure determination. MeshAndCollect can be applied to any type of micro-crystals, even if weakly diffracting. As an example, we present here the use of the technique to solve the crystal structure of the Cyan Fluorescent Protein (CFP) Cerulean.
X射线晶体学是用于获取有关生物大分子三维结构的高分辨率信息的主要技术。直到最近,一个主要要求是要有相对较大且衍射良好的晶体,而这些晶体往往很难获得。然而,串行晶体学的出现以及多晶体数据收集方法的复兴意味着大晶体的可用性不再是一个限制因素。在这里,我们展示了自动MeshAndCollect协议的使用,该协议首先识别安装在同一样品架上的许多小晶体的位置,然后指导从一系列部分衍射数据集的晶体中收集数据,以便后续合并并用于结构测定。MeshAndCollect可应用于任何类型的微晶,即使衍射较弱。作为一个例子,我们在此展示了使用该技术解析青色荧光蛋白(CFP)天蓝蛋白的晶体结构。