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Title: On-chip crystallization for serial crystallography experiments and on-chip ligand-binding studies

Abstract

Efficient and reliable sample delivery has remained one of the bottlenecks for serial crystallography experiments. Compared with other methods, fixed-target sample delivery offers the advantage of significantly reduced sample consumption and shorter data collection times owing to higher hit rates. Here, a new method of on-chip crystallization is reported which allows the efficient and reproducible growth of large numbers of protein crystals directly on micro-patterned silicon chips forin-situserial crystallography experiments. Crystals are grown by sitting-drop vapor diffusion and previously established crystallization conditions can be directly applied. By reducing the number of crystal-handling steps, the method is particularly well suited for sensitive crystal systems. Excessive mother liquor can be efficiently removed from the crystals by blotting, and no sealing of the fixed-target sample holders is required to prevent the crystals from dehydrating. As a consequence, `naked' crystals are obtained on the chip, resulting in very low background scattering levels and making the crystals highly accessible for external manipulation such as the application of ligand solutions. Serial diffraction experiments carried out at cryogenic temperatures at a synchrotron and at room temperature at an X-ray free-electron laser yielded high-quality X-ray structures of the human membrane protein aquaporin 2 and two new ligand-bound structuresmore » of thermolysin and the human kinase DRAK2. The results highlight the applicability of the method for future high-throughput on-chip screening of pharmaceutical compounds.« less

Authors:
ORCiD logo [1];  [1];  [2];  [1];  [1];  [1];  [1];  [1];  [1]; ORCiD logo [1];  [3]; ORCiD logo [4];  [1];  [1]; ORCiD logo [1];  [5];  [5];  [1];  [1];  [2] more »; ORCiD logo [6]; ORCiD logo [1] « less
  1. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany)
  2. Lund Univ. (Sweden)
  3. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); EMBL, Hamburg (Germany)
  4. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); Max Planck Institute of Biochemistry (Germany)
  5. SLAC National Accelerator Lab., Menlo Park, CA (United States)
  6. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); Univ. of Hamburg (Germany)
Publication Date:
Research Org.:
SLAC National Accelerator Lab., Menlo Park, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22)
OSTI Identifier:
1546938
Grant/Contract Number:  
AC02-76SF00515; 654220; 609920; 05K2018 – 2017-06727 MXD; 2010-5208; 2012-2849
Resource Type:
Accepted Manuscript
Journal Name:
IUCrJ
Additional Journal Information:
Journal Volume: 6; Journal Issue: 4; Journal ID: ISSN 2052-2525
Publisher:
International Union of Crystallography
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; serial crystallography; silicon chip; fixed-target crystallography; in-situ diffraction; vapor diffusion; ligand binding; ligand soaking; drug discovery; protein structure; X-ray crystallography

Citation Formats

Lieske, Julia, Cerv, Maximilian, Kreida, Stefan, Komadina, Dana, Fischer, Janine, Barthelmess, Miriam, Fischer, Pontus, Pakendorf, Tim, Yefanov, Oleksandr, Mariani, Valerio, Seine, Thomas, Ross, Breyan H., Crosas, Eva, Lorbeer, Olga, Burkhardt, Anja, Lane, Thomas J., Guenther, Sebastian, Bergtholdt, Julian, Schoen, Silvan, Törnroth-Horsefield, Susanna, Chapman, Henry N., and Meents, Alke. On-chip crystallization for serial crystallography experiments and on-chip ligand-binding studies. United States: N. p., 2019. Web. doi:10.1107/s2052252519007395.
Lieske, Julia, Cerv, Maximilian, Kreida, Stefan, Komadina, Dana, Fischer, Janine, Barthelmess, Miriam, Fischer, Pontus, Pakendorf, Tim, Yefanov, Oleksandr, Mariani, Valerio, Seine, Thomas, Ross, Breyan H., Crosas, Eva, Lorbeer, Olga, Burkhardt, Anja, Lane, Thomas J., Guenther, Sebastian, Bergtholdt, Julian, Schoen, Silvan, Törnroth-Horsefield, Susanna, Chapman, Henry N., & Meents, Alke. On-chip crystallization for serial crystallography experiments and on-chip ligand-binding studies. United States. doi:10.1107/s2052252519007395.
Lieske, Julia, Cerv, Maximilian, Kreida, Stefan, Komadina, Dana, Fischer, Janine, Barthelmess, Miriam, Fischer, Pontus, Pakendorf, Tim, Yefanov, Oleksandr, Mariani, Valerio, Seine, Thomas, Ross, Breyan H., Crosas, Eva, Lorbeer, Olga, Burkhardt, Anja, Lane, Thomas J., Guenther, Sebastian, Bergtholdt, Julian, Schoen, Silvan, Törnroth-Horsefield, Susanna, Chapman, Henry N., and Meents, Alke. Wed . "On-chip crystallization for serial crystallography experiments and on-chip ligand-binding studies". United States. doi:10.1107/s2052252519007395. https://www.osti.gov/servlets/purl/1546938.
@article{osti_1546938,
title = {On-chip crystallization for serial crystallography experiments and on-chip ligand-binding studies},
author = {Lieske, Julia and Cerv, Maximilian and Kreida, Stefan and Komadina, Dana and Fischer, Janine and Barthelmess, Miriam and Fischer, Pontus and Pakendorf, Tim and Yefanov, Oleksandr and Mariani, Valerio and Seine, Thomas and Ross, Breyan H. and Crosas, Eva and Lorbeer, Olga and Burkhardt, Anja and Lane, Thomas J. and Guenther, Sebastian and Bergtholdt, Julian and Schoen, Silvan and Törnroth-Horsefield, Susanna and Chapman, Henry N. and Meents, Alke},
abstractNote = {Efficient and reliable sample delivery has remained one of the bottlenecks for serial crystallography experiments. Compared with other methods, fixed-target sample delivery offers the advantage of significantly reduced sample consumption and shorter data collection times owing to higher hit rates. Here, a new method of on-chip crystallization is reported which allows the efficient and reproducible growth of large numbers of protein crystals directly on micro-patterned silicon chips forin-situserial crystallography experiments. Crystals are grown by sitting-drop vapor diffusion and previously established crystallization conditions can be directly applied. By reducing the number of crystal-handling steps, the method is particularly well suited for sensitive crystal systems. Excessive mother liquor can be efficiently removed from the crystals by blotting, and no sealing of the fixed-target sample holders is required to prevent the crystals from dehydrating. As a consequence, `naked' crystals are obtained on the chip, resulting in very low background scattering levels and making the crystals highly accessible for external manipulation such as the application of ligand solutions. Serial diffraction experiments carried out at cryogenic temperatures at a synchrotron and at room temperature at an X-ray free-electron laser yielded high-quality X-ray structures of the human membrane protein aquaporin 2 and two new ligand-bound structures of thermolysin and the human kinase DRAK2. The results highlight the applicability of the method for future high-throughput on-chip screening of pharmaceutical compounds.},
doi = {10.1107/s2052252519007395},
journal = {IUCrJ},
number = 4,
volume = 6,
place = {United States},
year = {2019},
month = {6}
}

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