Université Grenoble Alpes/CNRS/CEA/INRA/BIG

4 ARTICLES PUBLISHED IN JoVE

Biochemistry

Online Size-exclusion and Ion-exchange Chromatography on a SAXS Beamline

Martha E. Brennich¹, Adam R. Round^2,3, Stephanie Hutin⁴

¹Structural Biology Group, European Synchrotron Radiation Facility, ²European Molecular Biology Laboratory, ³School of Chemical and Physical Sciences, Keele University, ⁴Groupe de Microscopie Electronique et Méthodes, Institut de Biologie Structurale

The determination of the solution structure of a protein by small angle X-ray scattering (SAXS) requires monodisperse samples. Here, we present two possibilities to ensure minimal delays between sample preparation and data acquisition: online size-exclusion chromatography (SEC) and online ion-exchange chromatography (IEC).

Biochemistry

Structure Solution of the Fluorescent Protein Cerulean Using MeshAndCollect

Stephanie Hutin^*1, Gianluca Santoni^*1, Ulrich Zander², Nicolas Foos¹, Sylvain Aumonier¹, Guillaume Gotthard¹, Antoine Royant^1,3, Christoph Mueller-Dieckmann¹, Gordon Leonard¹

¹European Synchrotron Radiation Facility, Structural Biology Group, ²European Molecular Biology Laboratory, ³Univ. Grenoble Alpes, CNRS, CEA, IBS (Institut de Biologie Structurale)

We present the use of the MeshAndCollect protocol to obtain a complete diffraction data set, for use in subsequent structure determination, composed of partial diffraction data sets collected from many small crystals of the fluorescent protein Cerulean.

Biochemistry

Fully Autonomous Characterization and Data Collection from Crystals of Biological Macromolecules

Stephanie Hutin¹, Bart Van Laer¹, Christoph Mueller-Dieckmann¹, Gordon Leonard¹, Didier Nurizzo¹, Matthew W. Bowler²

¹Structural Biology Group, European Synchrotron Radiation Facility, ²Grenoble Outstation, European Molecular Biology Laboratory

Here, we describe how to use the automated screening and data collection options available at some synchrotron beamlines. Scientists send cryocooled samples to the synchrotron, and the diffraction properties are screened, the data sets are collected and processed and, where possible, a structure solution is carried out—all without human intervention.

Biochemistry

Analysis of SEC-SAXS data via EFA deconvolution and Scatter

Mark D Tully^*1, Nicolas Tarbouriech², Robert P Rambo³, Stephanie Hutin^*4

¹European Synchrotron Radiation Facility Structural Biology Group, Structural Biology Group, ²Institut de Biologie Structurale, University Grenoble Alpes, CEA, CNRS, ³Diamond Light Source, ⁴Laboratoire de Physiologie Cellulaire and Végétale, Université Grenoble Alpes/CNRS/CEA/INRA/BIG

SEC-BioSAXS measurements of biological macromolecules are a standard approach for determining solution structure of macromolecules and their complexes. Here, we analyze SEC-BioSAXS data from two types of commonly encountered SEC traces—chromatograms with fully resolved and partially resolved peaks. We demonstrate the analysis and deconvolution using scatter and BioXTAS RAW.