IUCrJ (Jul 2015)
A novel inert crystal delivery medium for serial femtosecond crystallography
- Chelsie E. Conrad,
- Shibom Basu,
- Daniel James,
- Dingjie Wang,
- Alexander Schaffer,
- Shatabdi Roy-Chowdhury,
- Nadia A. Zatsepin,
- Andrew Aquila,
- Jesse Coe,
- Cornelius Gati,
- Mark S. Hunter,
- Jason E. Koglin,
- Christopher Kupitz,
- Garrett Nelson,
- Ganesh Subramanian,
- Thomas A. White,
- Yun Zhao,
- James Zook,
- Sébastien Boutet,
- Vadim Cherezov,
- John C. H. Spence,
- Raimund Fromme,
- Uwe Weierstall,
- Petra Fromme
Affiliations
- Chelsie E. Conrad
- Department of Chemistry and Biochemistry, Arizona State University, PO Box 871604, Tempe, AZ 85287-1604, USA
- Shibom Basu
- Department of Chemistry and Biochemistry, Arizona State University, PO Box 871604, Tempe, AZ 85287-1604, USA
- Daniel James
- Center for Applied Structural Discovery, The Biodesign Institute, PO Box 875001, Tempe, AZ 85287-5001, USA
- Dingjie Wang
- Department of Physics, Arizona State University, PO Box 871604, Tempe, AZ 85287-1504, USA
- Alexander Schaffer
- Department of Chemistry and Biochemistry, Arizona State University, PO Box 871604, Tempe, AZ 85287-1604, USA
- Shatabdi Roy-Chowdhury
- Department of Chemistry and Biochemistry, Arizona State University, PO Box 871604, Tempe, AZ 85287-1604, USA
- Nadia A. Zatsepin
- Center for Applied Structural Discovery, The Biodesign Institute, PO Box 875001, Tempe, AZ 85287-5001, USA
- Andrew Aquila
- SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, CA 94025, USA
- Jesse Coe
- Department of Chemistry and Biochemistry, Arizona State University, PO Box 871604, Tempe, AZ 85287-1604, USA
- Cornelius Gati
- Center for Free-Electron Laser Science, Deutsches Elektronen-Synchrotron DESY, Notkestrasse 85, 22607 Hamburg, Germany
- Mark S. Hunter
- SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, CA 94025, USA
- Jason E. Koglin
- SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, CA 94025, USA
- Christopher Kupitz
- Center for Applied Structural Discovery, The Biodesign Institute, PO Box 875001, Tempe, AZ 85287-5001, USA
- Garrett Nelson
- Center for Applied Structural Discovery, The Biodesign Institute, PO Box 875001, Tempe, AZ 85287-5001, USA
- Ganesh Subramanian
- Center for Applied Structural Discovery, The Biodesign Institute, PO Box 875001, Tempe, AZ 85287-5001, USA
- Thomas A. White
- Center for Free-Electron Laser Science, Deutsches Elektronen-Synchrotron DESY, Notkestrasse 85, 22607 Hamburg, Germany
- Yun Zhao
- Center for Applied Structural Discovery, The Biodesign Institute, PO Box 875001, Tempe, AZ 85287-5001, USA
- James Zook
- Department of Chemistry and Biochemistry, Arizona State University, PO Box 871604, Tempe, AZ 85287-1604, USA
- Sébastien Boutet
- SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, CA 94025, USA
- Vadim Cherezov
- Bridge Institute, Department of Chemistry, University of Southern California, 3430 S. Vermont Avenue, Los Angeles, CA 90089, USA
- John C. H. Spence
- Center for Applied Structural Discovery, The Biodesign Institute, PO Box 875001, Tempe, AZ 85287-5001, USA
- Raimund Fromme
- Department of Chemistry and Biochemistry, Arizona State University, PO Box 871604, Tempe, AZ 85287-1604, USA
- Uwe Weierstall
- Center for Applied Structural Discovery, The Biodesign Institute, PO Box 875001, Tempe, AZ 85287-5001, USA
- Petra Fromme
- Department of Chemistry and Biochemistry, Arizona State University, PO Box 871604, Tempe, AZ 85287-1604, USA
- DOI
- https://doi.org/10.1107/S2052252515009811
- Journal volume & issue
-
Vol. 2,
no. 4
pp. 421 – 430
Abstract
Serial femtosecond crystallography (SFX) has opened a new era in crystallography 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.
Keywords
- serial femtosecond crystallography
- viscous crystal delivery
- protein complexes
- membrane proteins
- femtosecond studies
- nanocrystals
- coherent X-ray diffractive imaging
- free-electron laser
