title: A novel inert crystal delivery medium for serial femtosecond crystallography.
abstract: Serial femtosecond crystallography (SFX) has opened a new era in crystallo-graphy by permitting nearly damage-free, room-temperature structure determination of challenging proteins such as membrane proteins. In SFX, femtosecond X-ray free-electron laser pulses produce diffraction snapshots from nanocrystals and microcrystals delivered in a liquid jet, which leads to high protein consumption. A slow-moving stream of agarose has been developed as a new crystal delivery medium for SFX. It has low background scattering, is compatible with both soluble and membrane proteins, and can deliver the protein crystals at a wide range of temperatures down to 4°C. Using this crystal-laden agarose stream, the structure of a multi-subunit complex, phycocyanin, was solved to 2.5 Å resolution using 300 µg of microcrystals embedded into the agarose medium post-crystallization. The agarose delivery method reduces protein consumption by at least 100-fold and has the potential to be used for a diverse population of proteins, including membrane protein complexes.
authors.affiliation: Department of Chemistry and Biochemistry, Arizona State University , PO Box 871604, Tempe, AZ 85287-1604, USA ; Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA.
authors.forename: Chelsie E
authors.initials: CE
authors.lastname: Conrad
authors.affiliation: Department of Chemistry and Biochemistry, Arizona State University , PO Box 871604, Tempe, AZ 85287-1604, USA ; Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA.
authors.forename: Shibom
authors.initials: S
authors.lastname: Basu
authors.affiliation: Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA ; Department of Physics, Arizona State University , PO Box 871604, Tempe, AZ 85287-1504, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: James
authors.affiliation: Department of Physics, Arizona State University , PO Box 871604, Tempe, AZ 85287-1504, USA.
authors.forename: Dingjie
authors.initials: D
authors.lastname: Wang
authors.affiliation: Department of Chemistry and Biochemistry, Arizona State University , PO Box 871604, Tempe, AZ 85287-1604, USA ; Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA.
authors.forename: Alexander
authors.initials: A
authors.lastname: Schaffer
authors.affiliation: Department of Chemistry and Biochemistry, Arizona State University , PO Box 871604, Tempe, AZ 85287-1604, USA ; Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA.
authors.forename: Shatabdi
authors.initials: S
authors.lastname: Roy-Chowdhury
authors.affiliation: Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA ; Department of Physics, Arizona State University , PO Box 871604, Tempe, AZ 85287-1504, USA.
authors.forename: Nadia A
authors.initials: NA
authors.lastname: Zatsepin
authors.affiliation: SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, CA 94025, USA.
authors.forename: Andrew
authors.initials: A
authors.lastname: Aquila
authors.affiliation: Department of Chemistry and Biochemistry, Arizona State University , PO Box 871604, Tempe, AZ 85287-1604, USA ; Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA.
authors.forename: Jesse
authors.initials: J
authors.lastname: Coe
authors.affiliation: Center for Free-Electron Laser Science, Deutsches Elektronen-Synchrotron DESY , Notkestrasse 85, 22607 Hamburg, Germany.
authors.forename: Cornelius
authors.initials: C
authors.lastname: Gati
authors.affiliation: SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, CA 94025, USA.
authors.forename: Mark S
authors.initials: MS
authors.lastname: Hunter
authors.affiliation: SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, CA 94025, USA.
authors.forename: Jason E
authors.initials: JE
authors.lastname: Koglin
authors.affiliation: Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA ; Department of Physics, University of Wisconsin-Milwaukee , 1900 East Kenwood Boulevard, Milwaukee, WI 53211, USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: Kupitz
authors.affiliation: Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA ; Department of Physics, Arizona State University , PO Box 871604, Tempe, AZ 85287-1504, USA.
authors.forename: Garrett
authors.initials: G
authors.lastname: Nelson
authors.affiliation: Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA ; Department of Physics, Arizona State University , PO Box 871604, Tempe, AZ 85287-1504, USA.
authors.forename: Ganesh
authors.initials: G
authors.lastname: Subramanian
authors.affiliation: Center for Free-Electron Laser Science, Deutsches Elektronen-Synchrotron DESY , Notkestrasse 85, 22607 Hamburg, Germany.
authors.forename: Thomas A
authors.initials: TA
authors.lastname: White
authors.affiliation: Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA ; Department of Physics, Arizona State University , PO Box 871604, Tempe, AZ 85287-1504, USA.
authors.forename: Yun
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: Department of Chemistry and Biochemistry, Arizona State University , PO Box 871604, Tempe, AZ 85287-1604, USA ; Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA.
authors.forename: James
authors.initials: J
authors.lastname: Zook
authors.affiliation: SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, CA 94025, USA.
authors.forename: Sébastien
authors.initials: S
authors.lastname: Boutet
authors.affiliation: Bridge Institute, Department of Chemistry, University of Southern California , 3430 S. Vermont Avenue, Los Angeles, CA 90089, USA.
authors.forename: Vadim
authors.initials: V
authors.lastname: Cherezov
authors.affiliation: Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA ; Department of Physics, Arizona State University , PO Box 871604, Tempe, AZ 85287-1504, USA.
authors.forename: John C H
authors.initials: JC
authors.lastname: Spence
authors.affiliation: Department of Chemistry and Biochemistry, Arizona State University , PO Box 871604, Tempe, AZ 85287-1604, USA ; Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA.
authors.forename: Raimund
authors.initials: R
authors.lastname: Fromme
authors.affiliation: Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA ; Department of Physics, Arizona State University , PO Box 871604, Tempe, AZ 85287-1504, USA.
authors.forename: Uwe
authors.initials: U
authors.lastname: Weierstall
authors.affiliation: Department of Chemistry and Biochemistry, Arizona State University , PO Box 871604, Tempe, AZ 85287-1604, USA ; Center for Applied Structural Discovery, The Biodesign Institute , PO Box 875001, Tempe, AZ 85287-5001, USA.
authors.forename: Petra
authors.initials: P
authors.lastname: Fromme
keywords: coherent X-ray diffractive imaging; femtosecond studies; free-electron laser; membrane proteins; nanocrystals; protein complexes; serial femtosecond crystallography; viscous crystal delivery