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Title: Structure determination of an integral membrane protein at room temperature from crystals in situ

Abstract

The X-ray structure determination of an integral membrane protein using synchrotron diffraction data measured in situ at room temperature is demonstrated. The structure determination of an integral membrane protein using synchrotron X-ray diffraction data collected at room temperature directly in vapour-diffusion crystallization plates (in situ) is demonstrated. Exposing the crystals in situ eliminates manual sample handling and, since it is performed at room temperature, removes the complication of cryoprotection and potential structural anomalies induced by sample cryocooling. Essential to the method is the ability to limit radiation damage by recording a small amount of data per sample from many samples and subsequently assembling the resulting data sets using specialized software. The validity of this procedure is established by the structure determination of Haemophilus influenza TehA at 2.3 Å resolution. The method presented offers an effective protocol for the fast and efficient determination of membrane-protein structures at room temperature using third-generation synchrotron beamlines.

Authors:
 [1];  [1];  [2]; ;  [1];  [2];  [2];  [1];  [2];  [2];  [2];  [3];  [1];  [2];  [2];  [1];  [1];  [2];  [2]
  1. Diamond Light Source, Harwell Science and Innovation Campus, Oxfordshire OX11 0DE (United Kingdom)
  2. (United Kingdom)
  3. (Japan)
Publication Date:
OSTI Identifier:
22389067
Resource Type:
Journal Article
Journal Name:
Acta Crystallographica. Section D: Biological Crystallography
Additional Journal Information:
Journal Volume: 71; Journal Issue: Pt 6; Other Information: PMCID: PMC4461203; PMID: 26057664; PUBLISHER-ID: tz5076; OAI: oai:pubmedcentral.nih.gov:4461203; Copyright (c) Axford et al. 2015; This is an open-access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.; Country of input: International Atomic Energy Agency (IAEA); Journal ID: ISSN 0907-4449
Country of Publication:
Denmark
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; CRYSTALLIZATION; CRYSTALS; DIFFUSION; MEMBRANE PROTEINS; PLATES; POTENTIALS; RESOLUTION; SYNCHROTRONS; X-RAY DIFFRACTION

Citation Formats

Axford, Danny, Foadi, James, Imperial College London, London SW7 2AZ, Hu, Nien-Jen, Choudhury, Hassanul Ghani, Imperial College London, London SW7 2AZ, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, Iwata, So, Diamond Light Source, Harwell Science and Innovation Campus, Oxfordshire OX11 0DE, Imperial College London, London SW7 2AZ, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, Kyoto University, Kyoto 606-8501, Beis, Konstantinos, Imperial College London, London SW7 2AZ, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, Evans, Gwyndaf, E-mail: gwyndaf.evans@diamond.ac.uk, Alguel, Yilmaz, E-mail: gwyndaf.evans@diamond.ac.uk, Imperial College London, London SW7 2AZ, and Rutherford Appleton Laboratory, Oxfordshire OX11 0FA. Structure determination of an integral membrane protein at room temperature from crystals in situ. Denmark: N. p., 2015. Web. doi:10.1107/S139900471500423X.
Axford, Danny, Foadi, James, Imperial College London, London SW7 2AZ, Hu, Nien-Jen, Choudhury, Hassanul Ghani, Imperial College London, London SW7 2AZ, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, Iwata, So, Diamond Light Source, Harwell Science and Innovation Campus, Oxfordshire OX11 0DE, Imperial College London, London SW7 2AZ, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, Kyoto University, Kyoto 606-8501, Beis, Konstantinos, Imperial College London, London SW7 2AZ, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, Evans, Gwyndaf, E-mail: gwyndaf.evans@diamond.ac.uk, Alguel, Yilmaz, E-mail: gwyndaf.evans@diamond.ac.uk, Imperial College London, London SW7 2AZ, & Rutherford Appleton Laboratory, Oxfordshire OX11 0FA. Structure determination of an integral membrane protein at room temperature from crystals in situ. Denmark. doi:10.1107/S139900471500423X.
Axford, Danny, Foadi, James, Imperial College London, London SW7 2AZ, Hu, Nien-Jen, Choudhury, Hassanul Ghani, Imperial College London, London SW7 2AZ, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, Iwata, So, Diamond Light Source, Harwell Science and Innovation Campus, Oxfordshire OX11 0DE, Imperial College London, London SW7 2AZ, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, Kyoto University, Kyoto 606-8501, Beis, Konstantinos, Imperial College London, London SW7 2AZ, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, Evans, Gwyndaf, E-mail: gwyndaf.evans@diamond.ac.uk, Alguel, Yilmaz, E-mail: gwyndaf.evans@diamond.ac.uk, Imperial College London, London SW7 2AZ, and Rutherford Appleton Laboratory, Oxfordshire OX11 0FA. Thu . "Structure determination of an integral membrane protein at room temperature from crystals in situ". Denmark. doi:10.1107/S139900471500423X.
@article{osti_22389067,
title = {Structure determination of an integral membrane protein at room temperature from crystals in situ},
author = {Axford, Danny and Foadi, James and Imperial College London, London SW7 2AZ and Hu, Nien-Jen and Choudhury, Hassanul Ghani and Imperial College London, London SW7 2AZ and Rutherford Appleton Laboratory, Oxfordshire OX11 0FA and Iwata, So and Diamond Light Source, Harwell Science and Innovation Campus, Oxfordshire OX11 0DE and Imperial College London, London SW7 2AZ and Rutherford Appleton Laboratory, Oxfordshire OX11 0FA and Kyoto University, Kyoto 606-8501 and Beis, Konstantinos and Imperial College London, London SW7 2AZ and Rutherford Appleton Laboratory, Oxfordshire OX11 0FA and Evans, Gwyndaf, E-mail: gwyndaf.evans@diamond.ac.uk and Alguel, Yilmaz, E-mail: gwyndaf.evans@diamond.ac.uk and Imperial College London, London SW7 2AZ and Rutherford Appleton Laboratory, Oxfordshire OX11 0FA},
abstractNote = {The X-ray structure determination of an integral membrane protein using synchrotron diffraction data measured in situ at room temperature is demonstrated. The structure determination of an integral membrane protein using synchrotron X-ray diffraction data collected at room temperature directly in vapour-diffusion crystallization plates (in situ) is demonstrated. Exposing the crystals in situ eliminates manual sample handling and, since it is performed at room temperature, removes the complication of cryoprotection and potential structural anomalies induced by sample cryocooling. Essential to the method is the ability to limit radiation damage by recording a small amount of data per sample from many samples and subsequently assembling the resulting data sets using specialized software. The validity of this procedure is established by the structure determination of Haemophilus influenza TehA at 2.3 Å resolution. The method presented offers an effective protocol for the fast and efficient determination of membrane-protein structures at room temperature using third-generation synchrotron beamlines.},
doi = {10.1107/S139900471500423X},
journal = {Acta Crystallographica. Section D: Biological Crystallography},
issn = {0907-4449},
number = Pt 6,
volume = 71,
place = {Denmark},
year = {2015},
month = {5}
}