title: Drop-on-demand sample delivery for studying biocatalysts in action at X-ray free-electron lasers.
abstract: X-ray crystallography at X-ray free-electron laser sources is a powerful method for studying macromolecules at biologically relevant temperatures. Moreover, when combined with complementary techniques like X-ray emission spectroscopy, both global structures and chemical properties of metalloenzymes can be obtained concurrently, providing insights into the interplay between the protein structure and dynamics and the chemistry at an active site. The implementation of such a multimodal approach can be compromised by conflicting requirements to optimize each individual method. In particular, the method used for sample delivery greatly affects the data quality. We present here a robust way of delivering controlled sample amounts on demand using acoustic droplet ejection coupled with a conveyor belt drive that is optimized for crystallography and spectroscopy measurements of photochemical and chemical reactions over a wide range of time scales. Studies with photosystem II, the phytochrome photoreceptor, and ribonucleotide reductase R2 illustrate the power and versatility of this method.
authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.affiliation: Department of Biology, Washington University in St. Louis, St. Louis, Missouri, USA.
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.affiliation: Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden.
authors.forename: Hugo
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authors.affiliation: Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden.
authors.forename: Vivek
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.lastname: Michels-Clark
authors.affiliation: Department of BioSciences, Rice University, Houston, Texas, USA.
authors.forename: Jonathan A
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authors.forename: Babak
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authors.lastname: Andi
authors.affiliation: Institut für Biologie, Humboldt-Universität zu Berlin, Berlin, Germany.
authors.forename: Mohamed
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.forename: Casper
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authors.affiliation: Institut für Biologie, Humboldt-Universität zu Berlin, Berlin, Germany.
authors.forename: Rana
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authors.affiliation: Department of Chemistry, Pennsylvania State University, University Park, Pennsylvania, USA.
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authors.affiliation: Institut für Biologie, Humboldt-Universität zu Berlin, Berlin, Germany.
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authors.affiliation: SSRL, SLAC National Accelerator Laboratory, Menlo Park, California, USA.
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authors.affiliation: Stanford PULSE Institute, SLAC National Accelerator Laboratory, Menlo Park, California, USA.
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authors.lastname: Kubin
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.affiliation: Diamond Light Source Limited, Harwell Science and Innovation Campus, Didcot, UK.|Department of Biochemistry, University of Oxford, Oxford, UK.
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authors.lastname: Bräuer
authors.affiliation: Department of BioSciences, Rice University, Houston, Texas, USA.
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.affiliation: Institutionen för Kemi, Kemiskt Biologiskt Centrum, Umeå Universitet, Umeå, Sweden.|Stanford PULSE Institute, SLAC National Accelerator Laboratory, Menlo Park, California, USA.
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authors.affiliation: National Synchrotron Light Source II, Brookhaven National Laboratory, Upton, New York, USA.
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.identifier: http://orcid.org/0000-0002-6253-8282
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authors.lastname: Sierra
authors.affiliation: Diamond Light Source Limited, Harwell Science and Innovation Campus, Didcot, UK.
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authors.affiliation: LCLS, SLAC National Accelerator Laboratory, Menlo Park, California, USA.
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authors.affiliation: LCLS, SLAC National Accelerator Laboratory, Menlo Park, California, USA.
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authors.affiliation: LCLS, SLAC National Accelerator Laboratory, Menlo Park, California, USA.
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authors.identifier: http://orcid.org/0000-0001-6811-6083
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authors.affiliation: LCLS, SLAC National Accelerator Laboratory, Menlo Park, California, USA.
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authors.affiliation: LCLS, SLAC National Accelerator Laboratory, Menlo Park, California, USA.
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authors.affiliation: Institutionen för Kemi, Kemiskt Biologiskt Centrum, Umeå Universitet, Umeå, Sweden.|Department of Chemistry-Ångström, Molecular Biomimetics, Uppsala University, Uppsala, Sweden.
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authors.affiliation: Department of BioSciences, Rice University, Houston, Texas, USA.|Department of Chemistry, Rice University, Houston, Texas, USA.
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authors.affiliation: Department of Biology, Washington University in St. Louis, St. Louis, Missouri, USA.
authors.forename: Richard D
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
authors.forename: Nicholas K
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authors.affiliation: Diamond Light Source Limited, Harwell Science and Innovation Campus, Didcot, UK.
authors.forename: Allen M
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.|LCLS, SLAC National Accelerator Laboratory, Menlo Park, California, USA.
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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authors.affiliation: Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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mesh_terms: D000162:Acoustics; D018360:Crystallography, X-Ray / Q000379:methods*; D007834:Lasers; D045332:Photosystem II Protein Complex / Q000737:chemistry; D010834:Phytochrome / Q000737:chemistry; D012264:Ribonucleotide Reductases / Q000737:chemistry; D013052:Spectrometry, X-Ray Emission / Q000379:methods
chemical_list: D045332:Photosystem II Protein Complex; D010834:Phytochrome; D012264:Ribonucleotide Reductases