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Title: Double-flow focused liquid injector for efficient serial femtosecond crystallography

Abstract

Serial femtosecond crystallography requires reliable and efficient delivery of fresh crystals across the beam of an X-ray free-electron laser over the course of an experiment. We introduce a double-flow focusing nozzle to meet this challenge, with significantly reduced sample consumption, while improving jet stability over previous generations of nozzles. We demonstrate its use to determine the first room-temperature structure of RNA polymerase II at high resolution, revealing new structural details. Furthermore, the double flow-focusing nozzles were successfully tested with three other protein samples and the first room temperature structure of an extradiol ring-cleaving dioxygenase was solved by utilizing the improved operation and characteristics of these devices.

Authors:
 [1];  [2];  [2];  [1];  [3];  [4];  [5];  [1];  [6];  [1];  [1];  [2];  [1];  [7];  [1];  [8];  [9];  [1];  [10];  [6] more »;  [6];  [11];  [11];  [12];  [12];  [13];  [13];  [14];  [4];  [15];  [16];  [16];  [17];  [4];  [4];  [4];  [4];  [12];  [18];  [6];  [3];  [6];  [11];  [19];  [1] « less
  1. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany)
  2. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); Univ. of Hamburg, Hamburg (Germany)
  3. Stanford Univ., Stanford, CA (United States)
  4. SLAC National Accelerator Lab., Menlo Park, CA (United States)
  5. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); Max-Planck-Institute for Biochemistry, Munich (Germany)
  6. Arizona State Univ., Tempe, AZ (United States)
  7. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); Centre for Ultrafast Imaging, Hamburg (Germany)
  8. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); European XFEL GmbH, Hamburg (Germany)
  9. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); Univ. of Hamburg, Hamburg (Germany); Max-Planck Institute for the Structure and Dynamics of Matter, Hamburg (Germany)
  10. Arizona State Univ., Tempe, AZ (United States); Paul Scherrer Inst. (PSI), Villigen (Switzerland)
  11. Cornell Univ., Ithaca, NY (United States)
  12. Charite - Univ. Berlin, Berlin (Germany)
  13. Technische Univ. Berlin, Berlin (Germany)
  14. Hauptman-Woodward Medical Research Institute, Buffalo, NY (United States)
  15. Univ. of Nova Gorica, Nova Gorica (Slovenia); Institute of Metals and Technology, Ljubljana (Slovenia)
  16. Institute of Metals and Technology, Ljubljana (Slovenia)
  17. Helmholtz-Zentrum Geesthacht, Geesthacht (Germany)
  18. Univ. of Minnesota, Minneapolis, MN (United States)
  19. Deutsches Elektronen-Synchrotron (DESY), Hamburg (Germany); Univ. of Hamburg, Hamburg (Germany); Univ. of Minnesota, Minneapolis, MN (United States)
Publication Date:
Research Org.:
SLAC National Accelerator Lab., Menlo Park, CA (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1352549
Grant/Contract Number:
AC02-76SF00515
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Scientific Reports
Additional Journal Information:
Journal Volume: 7; Journal ID: ISSN 2045-2322
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
English
Subject:
71 CLASSICAL AND QUANTUM MECHANICS, GENERAL PHYSICS; fluid dynamics; nanocrystallography

Citation Formats

Oberthuer, Dominik, Knoška, Juraj, Wiedorn, Max O., Beyerlein, Kenneth R., Bushnell, David A., Kovaleva, Elena G., Heymann, Michael, Gumprecht, Lars, Kirian, Richard A., Barty, Anton, Mariani, Valerio, Tolstikova, Aleksandra, Adriano, Luigi, Awel, Salah, Barthelmess, Miriam, Dörner, Katerina, Xavier, P. Lourdu, Yefanov, Oleksandr, James, Daniel R., Nelson, Garrett, Wang, Dingjie, Calvey, George, Chen, Yujie, Schmidt, Andrea, Szczepek, Michael, Frielingsdorf, Stefan, Lenz, Oliver, Snell, Edward, Robinson, Philip J., Šarler, Božidar, Belšak, Grega, Maček, Marjan, Wilde, Fabian, Aquila, Andrew, Boutet, Sébastien, Liang, Mengning, Hunter, Mark S., Scheerer, Patrick, Lipscomb, John D., Weierstall, Uwe, Kornberg, Roger D., Spence, John C. H., Pollack, Lois, Chapman, Henry N., and Bajt, Saša. Double-flow focused liquid injector for efficient serial femtosecond crystallography. United States: N. p., 2017. Web. doi:10.1038/srep44628.
Oberthuer, Dominik, Knoška, Juraj, Wiedorn, Max O., Beyerlein, Kenneth R., Bushnell, David A., Kovaleva, Elena G., Heymann, Michael, Gumprecht, Lars, Kirian, Richard A., Barty, Anton, Mariani, Valerio, Tolstikova, Aleksandra, Adriano, Luigi, Awel, Salah, Barthelmess, Miriam, Dörner, Katerina, Xavier, P. Lourdu, Yefanov, Oleksandr, James, Daniel R., Nelson, Garrett, Wang, Dingjie, Calvey, George, Chen, Yujie, Schmidt, Andrea, Szczepek, Michael, Frielingsdorf, Stefan, Lenz, Oliver, Snell, Edward, Robinson, Philip J., Šarler, Božidar, Belšak, Grega, Maček, Marjan, Wilde, Fabian, Aquila, Andrew, Boutet, Sébastien, Liang, Mengning, Hunter, Mark S., Scheerer, Patrick, Lipscomb, John D., Weierstall, Uwe, Kornberg, Roger D., Spence, John C. H., Pollack, Lois, Chapman, Henry N., & Bajt, Saša. Double-flow focused liquid injector for efficient serial femtosecond crystallography. United States. doi:10.1038/srep44628.
Oberthuer, Dominik, Knoška, Juraj, Wiedorn, Max O., Beyerlein, Kenneth R., Bushnell, David A., Kovaleva, Elena G., Heymann, Michael, Gumprecht, Lars, Kirian, Richard A., Barty, Anton, Mariani, Valerio, Tolstikova, Aleksandra, Adriano, Luigi, Awel, Salah, Barthelmess, Miriam, Dörner, Katerina, Xavier, P. Lourdu, Yefanov, Oleksandr, James, Daniel R., Nelson, Garrett, Wang, Dingjie, Calvey, George, Chen, Yujie, Schmidt, Andrea, Szczepek, Michael, Frielingsdorf, Stefan, Lenz, Oliver, Snell, Edward, Robinson, Philip J., Šarler, Božidar, Belšak, Grega, Maček, Marjan, Wilde, Fabian, Aquila, Andrew, Boutet, Sébastien, Liang, Mengning, Hunter, Mark S., Scheerer, Patrick, Lipscomb, John D., Weierstall, Uwe, Kornberg, Roger D., Spence, John C. H., Pollack, Lois, Chapman, Henry N., and Bajt, Saša. Thu . "Double-flow focused liquid injector for efficient serial femtosecond crystallography". United States. doi:10.1038/srep44628. https://www.osti.gov/servlets/purl/1352549.
@article{osti_1352549,
title = {Double-flow focused liquid injector for efficient serial femtosecond crystallography},
author = {Oberthuer, Dominik and Knoška, Juraj and Wiedorn, Max O. and Beyerlein, Kenneth R. and Bushnell, David A. and Kovaleva, Elena G. and Heymann, Michael and Gumprecht, Lars and Kirian, Richard A. and Barty, Anton and Mariani, Valerio and Tolstikova, Aleksandra and Adriano, Luigi and Awel, Salah and Barthelmess, Miriam and Dörner, Katerina and Xavier, P. Lourdu and Yefanov, Oleksandr and James, Daniel R. and Nelson, Garrett and Wang, Dingjie and Calvey, George and Chen, Yujie and Schmidt, Andrea and Szczepek, Michael and Frielingsdorf, Stefan and Lenz, Oliver and Snell, Edward and Robinson, Philip J. and Šarler, Božidar and Belšak, Grega and Maček, Marjan and Wilde, Fabian and Aquila, Andrew and Boutet, Sébastien and Liang, Mengning and Hunter, Mark S. and Scheerer, Patrick and Lipscomb, John D. and Weierstall, Uwe and Kornberg, Roger D. and Spence, John C. H. and Pollack, Lois and Chapman, Henry N. and Bajt, Saša},
abstractNote = {Serial femtosecond crystallography requires reliable and efficient delivery of fresh crystals across the beam of an X-ray free-electron laser over the course of an experiment. We introduce a double-flow focusing nozzle to meet this challenge, with significantly reduced sample consumption, while improving jet stability over previous generations of nozzles. We demonstrate its use to determine the first room-temperature structure of RNA polymerase II at high resolution, revealing new structural details. Furthermore, the double flow-focusing nozzles were successfully tested with three other protein samples and the first room temperature structure of an extradiol ring-cleaving dioxygenase was solved by utilizing the improved operation and characteristics of these devices.},
doi = {10.1038/srep44628},
journal = {Scientific Reports},
number = ,
volume = 7,
place = {United States},
year = {Thu Mar 16 00:00:00 EDT 2017},
month = {Thu Mar 16 00:00:00 EDT 2017}
}

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Cited by: 13works
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  • Cited by 2
  • We describe a concentric-flow electrokinetic injector for efficiently delivering microcrystals for serial femtosecond X-ray crystallography analysis that enables studies of challenging biological systems in their unadulterated mother liquor. We used the injector to analyze microcrystals of Geobacillus stearothermophilus thermolysin (2.2-Å structure), Thermosynechococcus elongatus photosystem II (<3-Å diffraction) and Thermus thermophilus small ribosomal subunit bound to the antibiotic paromomycin at ambient temperature (3.4-Å structure).
  • In this paper, we describe a concentric-flow electrokinetic injector for efficiently delivering microcrystals for serial femtosecond X-ray crystallography analysis that enables studies of challenging biological systems in their unadulterated mother liquor. Finally, we used the injector to analyze microcrystals of Geobacillus stearothermophilus thermolysin (2.2-Å structure), Thermosynechococcus elongatus photosystem II (<3-Å diffraction) and Thermus thermophilus small ribosomal subunit bound to the antibiotic paromomycin at ambient temperature (3.4-Å structure).
  • A raster scanning serial protein crystallography approach is presented, that consumes as low ∼200–700 nl of sedimented crystals. New serial data pre-analysis software, NanoPeakCell, is introduced. High-resolution structural information was obtained from lysozyme microcrystals (20 µm in the largest dimension) using raster-scanning serial protein crystallography on micro- and nano-focused beamlines at the ESRF. Data were collected at room temperature (RT) from crystals sandwiched between two silicon nitride wafers, thereby preventing their drying, while limiting background scattering and sample consumption. In order to identify crystal hits, new multi-processing and GUI-driven Python-based pre-analysis software was developed, named NanoPeakCell, that was able tomore » read data from a variety of crystallographic image formats. Further data processing was carried out using CrystFEL, and the resultant structures were refined to 1.7 Å resolution. The data demonstrate the feasibility of RT raster-scanning serial micro- and nano-protein crystallography at synchrotrons and validate it as an alternative approach for the collection of high-resolution structural data from micro-sized crystals. Advantages of the proposed approach are its thriftiness, its handling-free nature, the reduced amount of sample required, the adjustable hit rate, the high indexing rate and the minimization of background scattering.« less