Author:
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
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.
Publisher
International Union of Crystallography (IUCr)
Subject
Condensed Matter Physics,General Materials Science,Biochemistry,General Chemistry
Cited by
126 articles.
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