A novel inert crystal delivery medium for serial femtosecond crystallography

• Published in: IUCrJ 2015-06, Vol.2 (4)
• Main Authors: 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
• Format: Article
• Language: English
• ISSN: 2052-2525; 2052-2525

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 2.5Å resolution using 300µg 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.