Zeng Lingxiao, Ding Wei, Hao Quan
School of Biomedical Sciences, University of Hong Kong, 21 Sassoon Road, Hong Kong.
Institute of Physics, Chinese Academy of Sciences, Beijing 100190, People's Republic of China.
IUCrJ. 2018 May 11;5(Pt 4):382-389. doi: 10.1107/S2052252518005857. eCollection 2018 Jul 1.
X-ray crystallography and cryo-electron microscopy (cryo-EM) are complementary techniques for structure determination. Crystallography usually reveals more detailed information, while cryo-EM is an extremely useful technique for studying large-sized macromolecules. As the gap between the resolution of crystallography and cryo-EM data narrows, the cryo-EM map of a macromolecule could serve as an initial model to solve the phase problem of crystal diffraction for high-resolution structure determination. is a procedure to utilize the low-resolution molecular shape for crystallographic phasing. The (Iterative Protein Crystal structure Automatic Solution) pipeline is an automatic direct-methods-aided dual-space iterative phasing and model-building procedure. When only an electron-density map is available as the starting point, is capable of generating a completed model from the phases of the input map automatically, without the requirement of an initial model. In this study, a hybrid method integrating X-ray crystallography with cryo-EM to help with structure determination is presented. With a cryo-EM map as the starting point, the workflow of the method involves three steps. (1) Cryo-EM map replacement: is utilized to find the correct translation and orientation of the cryo-EM map in the crystallographic unit cell and generates the initial low-resolution map. (2) Phase extension: the phases calculated from the correctly placed cryo-EM map are extended to high-resolution X-ray data by non-crystallographic symmetry averaging with . (3) Model building: is used to generate an initial model using the phase-extended map and perform model completion by iteration. Four cases (the lowest cryo-EM map resolution being 6.9 Å) have been tested for the general applicability of the hybrid method, and almost complete models have been generated for all test cases with reasonable /. The hybrid method therefore provides an automated tool for X-ray structure determination using a cryo-EM map as the starting point.
X射线晶体学和冷冻电子显微镜技术（cryo-EM）是用于结构测定的互补技术。晶体学通常能揭示更详细的信息，而冷冻电子显微镜技术对于研究大型大分子是一项极其有用的技术。随着晶体学和冷冻电子显微镜数据分辨率之间的差距缩小，大分子的冷冻电子显微镜图谱可作为解决晶体衍射相位问题以进行高分辨率结构测定的初始模型。是一种利用低分辨率分子形状进行晶体学相位测定的方法。（迭代蛋白质晶体结构自动求解）流程是一种自动直接法辅助的双空间迭代相位测定和模型构建程序。当仅以电子密度图作为起点时，能够从输入图的相位自动生成完整模型，而无需初始模型。在本研究中，提出了一种将X射线晶体学与冷冻电子显微镜技术相结合以辅助结构测定的混合方法。以冷冻电子显微镜图谱为起点，该方法的工作流程包括三个步骤。（1）冷冻电子显微镜图谱替换：用于在晶体学晶胞中找到冷冻电子显微镜图谱的正确平移和取向，并生成初始低分辨率图谱。（2）相位扩展：通过与非晶体学对称平均将从正确放置的冷冻电子显微镜图谱计算出的相位扩展到高分辨率X射线数据。（3）模型构建：用于使用相位扩展后的图谱生成初始模型并通过迭代完成模型构建。已对四个案例（最低冷冻电子显微镜图谱分辨率为6.9 Å）测试了该混合方法的普遍适用性，并且在所有测试案例中都以合理的/生成了几乎完整的模型。因此，该混合方法提供了一种以冷冻电子显微镜图谱为起点进行X射线结构测定的自动化工具。
