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.
| Original language | English |
|---|---|
| Pages (from-to) | 421-430 |
| Number of pages | 10 |
| Journal | IUCrJ |
| Volume | 2 |
| DOIs | |
| Publication status | Published - Jul 1 2015 |
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Keywords
- coherent X-ray diffractive imaging
- femtosecond studies
- free-electron laser
- membrane proteins
- nanocrystals
- protein complexes
- serial femtosecond crystallography
- viscous crystal delivery
ASJC Scopus subject areas
- Chemistry(all)
- Materials Science(all)
- Condensed Matter Physics
- Biochemistry
Cite this
A novel inert crystal delivery medium for serial femtosecond crystallography. / Conrad, Chelsie E.; Basu, Shibom; James, Daniel; Wang, Dingjie; Schaffer, Alexander; Roy-Chowdhury, Shatabdi; Zatsepin, Nadia A.; Aquila, Andrew; Coe, Jesse; Gati, Cornelius; Hunter, Mark S.; Koglin, Jason E.; Kupitz, Christopher; Nelson, Garrett; Subramanian, Ganesh; White, Thomas A.; Zhao, Yun; Zook, James; Boutet, Sébastien; Cherezov, Vadim; Spence, John C H; Fromme, Raimund; Weierstall, Uwe; Fromme, Petra.
In: IUCrJ, Vol. 2, 01.07.2015, p. 421-430.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - A novel inert crystal delivery medium for serial femtosecond crystallography
AU - Conrad, Chelsie E.
AU - Basu, Shibom
AU - James, Daniel
AU - Wang, Dingjie
AU - Schaffer, Alexander
AU - Roy-Chowdhury, Shatabdi
AU - Zatsepin, Nadia A.
AU - Aquila, Andrew
AU - Coe, Jesse
AU - Gati, Cornelius
AU - Hunter, Mark S.
AU - Koglin, Jason E.
AU - Kupitz, Christopher
AU - Nelson, Garrett
AU - Subramanian, Ganesh
AU - White, Thomas A.
AU - Zhao, Yun
AU - Zook, James
AU - Boutet, Sébastien
AU - Cherezov, Vadim
AU - Spence, John C H
AU - Fromme, Raimund
AU - Weierstall, Uwe
AU - Fromme, Petra
PY - 2015/7/1
Y1 - 2015/7/1
N2 - 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.
AB - 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.
KW - coherent X-ray diffractive imaging
KW - femtosecond studies
KW - free-electron laser
KW - membrane proteins
KW - nanocrystals
KW - protein complexes
KW - serial femtosecond crystallography
KW - viscous crystal delivery
UR - http://www.scopus.com/inward/record.url?scp=84939784819&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84939784819&partnerID=8YFLogxK
U2 - 10.1107/S2052252515009811
DO - 10.1107/S2052252515009811
M3 - Article
VL - 2
SP - 421
EP - 430
JO - IUCrJ
JF - IUCrJ
SN - 2052-2525
ER -