De novo protein structure determination by heavy-atom soaking in lipidic cubic phase and SIRAS phasing using serial synchrotron crystallography

IUCrJ

Schmidt, C.¹  Botha, S.²  Baitan, D.³  Jungnickel, K.E.J.⁴  Oberthr, D.⁵ 

[1] Center for Free-Electron Laser Science, Deutsches Elektronen-Synchrotron (DESY), Notkestrasse 85, 22607 Hamburg, Germany;Institute of Biochemistry and Molecular Biology, Chemistry Department, University of Hamburg, Martin-Luther-King Platz 6, 20146 Hamburg, Germany;Laboratory for Structural Biology of Infection and Inflammation, c/o DESY, Building 22a, Notkestrasse 85, 22607 Hamburg, Germany;The Hamburg Centre for Ultrafast Imaging, Luruper Chaussee 149, 22761 Hamburg, Germany;Xtal Concepts GmbH, Marlowring 19, 22525 Hamburg, Germany

关键词: SERIAL CRYSTALLOGRAPHY;    SIRAS PHASING;    LIPIDIC CUBIC PHASE;    HEAVY-ATOM SOAKING;    DE NOVO PROTEIN STRUCTURE DETERMINATION;   

DOI  :  10.1107/S2052252518009223

学科分类：数学（综合）

来源: International Union of Crystallography

【 摘 要 】

During the past few years, serial crystallography methods have undergone continuous development and serial data collection has become well established at high-intensity synchrotron-radiation beamlines and XFEL radiation sources. However, the application of experimental phasing to serial crystallography data has remained a challenging task owing to the inherent inaccuracy of the diffraction data. Here, a particularly gentle method for incorporating heavy atoms into micrometre-sized crystals utilizing lipidic cubic phase (LCP) as a carrier medium is reported. Soaking in LCP prior to data collection offers a new, efficient and gentle approach for preparing heavy-atom-derivative crystals directly before diffraction data collection using serial crystallography methods. This approach supports effective phasing by utilizing a reasonably low number of diffraction patterns. Using synchrotron radiation and exploiting the anomalous scattering signal of mercury for single isomorphous replacement with anomalous scattering (SIRAS) phasing resulted in high-quality electron-density maps that were sufficient for building a complete structural model of proteinase K at 1.9 Å resolution using automatic model-building tools.

【 授权许可】

CC BY