Serial Femtosecond Crystallography of G Protein–Coupled Receptors

Liu, Wei; Wacker, Daniel; Gati, Cornelius; Han, Gye Won; James, Daniel; Wang, Dingjie; Nelson, Garrett; Weierstall, Uwe; Katritch, Vsevolod; Barty, Anton; Zatsepin, Nadia A.; Li, Dianfan; Messerschmidt, Marc; Boutet, Sébastien; Williams, Garth J.; Koglin, Jason E.; Seibert, M.; Wang, Chong; Shah, Syed Tasadaque Ali; Basu, Shibom; Fromme, Raimund; Kupitz, Christopher; Rendek, K.N.; Grotjohann, Ingo; Fromme, Petra; Kirian, Richard A.; Beyerlein, Kenneth R.; White, Thomas A.; Chapman, Henry N.; Caffrey, Martin; Spence, John C. H.; Stevens, Raymond C.; Cherezov, Vadim

doi:10.1126/science.1244142

Cited by 424 publications

(378 citation statements)

References 37 publications

(27 reference statements)

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“…In addition to the GDVN (2,3,14,16,17,19,20), other injectors such as a nanoflow electrospinning injector (21) and a lipidic cubic phase (LCP) injector (22), have been developed that have a reduced flow rate and lower sample consumption. However, because injectors deliver a continuous stream of solution containing a random distribution of crystals, and the X-ray pulses are extremely short, often only a small percentage of pulses hit a crystal and produce a useful diffraction pattern.…”

Section: Significancementioning

confidence: 99%

Goniometer-based femtosecond crystallography with X-ray free electron lasers

Cohen

Soltis

González

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

128

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The emerging method of femtosecond crystallography (FX) may extend the diffraction resolution accessible from small radiationsensitive crystals and provides a means to determine catalytically accurate structures of acutely radiation-sensitive metalloenzymes. Automated goniometer-based instrumentation developed for use at the Linac Coherent Light Source enabled efficient and flexible FX experiments to be performed on a variety of sample types. In the case of rod-shaped Cpl hydrogenase crystals, only five crystals and about 30 min of beam time were used to obtain the 125 still diffraction patterns used to produce a 1.6-Å resolution electron density map. For smaller crystals, high-density grids were used to increase sample throughput; 930 myoglobin crystals mounted at random orientation inside 32 grids were exposed, demonstrating the utility of this approach. Screening results from cryocooled crystals of β 2 -adrenoreceptor and an RNA polymerase II complex indicate the potential to extend the diffraction resolution obtainable from very radiation-sensitive samples beyond that possible with undulator-based synchrotron sources.femtosecond diffraction | crystallography | XFEL | structural biology U sing extremely bright, short-timescale X-ray pulses produced by X-ray free-electron lasers (XFELs), femtosecond crystallography (FX) is an emerging method that expands the structural information accessible from very small or very radiation-sensitive macromolecular crystals. Central to this method is the "diffraction before destruction" (1) process in which a still diffraction image is produced by a single X-ray pulse before significant radiation-induced electronic and atomic rearrangements occur within the crystal (1-3). At the Linac Coherent Light Source (LCLS) at SLAC, a single ∼50-fs-long X-ray pulse can expose a crystal to as many X-ray photons as a typical synchrotron beam line produces in about a second. Exposing small crystals to these intense ultrashort pulses circumvents the dose limitations of conventional X-ray diffraction experiments (4) and may produce useful data to resolutions beyond what is achievable at synchrotrons (5). This innovation provides a pathway to obtain atomic information from proteins that only form micrometer-to nanometer-sized crystals, such as many membrane proteins and large multiprotein complexes. Moreover, XFELs enable "diffraction before reduction" data collection to address another major challenge in structural enzymology by providing a means to determine catalytically accurate structures of acutely radiation-sensitive metalloenzyme active sites (6), such as high-valency reaction intermediates that may be significantly photoreduced during a single X-ray exposure at a synchrotron, even at very small doses (7-11). Furthermore, the use of short (tens of femtoseconds) X-ray pulses further complements the structural characterization of biochemical reaction processes by providing access to a time domain two to three orders of magnitude faster (12, 13) than currently accessible using synchrotro...

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Section: Significancementioning

confidence: 99%

Goniometer-based femtosecond crystallography with X-ray free electron lasers

Cohen

Soltis

González

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

128

View full text Add to dashboard Cite

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“…The technique merges thousands to tens of thousands of twodimensional diffraction patterns to obtain a three-dimensional data set which can then be analyzed using standard protein crystal phasing techniques. The first experiments were performed on well known model systems (Chapman et al, 2011;Boutet et al, 2012) and, since the establishment of SFX, it has been widely applied to systems or dynamics that were difficult or impossible to study at room temperature using more conventional macromolecular crystallography approaches (Kern et al, 2013; Kupitz et al, 2014;Liu et al, 2013;Demirci et al, 2013).…”

Section: Research Highlightsmentioning

confidence: 99%

The Coherent X-ray Imaging instrument at the Linac Coherent Light Source

et al. 2015

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The Coherent X-ray Imaging (CXI) instrument specializes in hard X-ray, invacuum, high power density experiments in all areas of science. Two main sample chambers, one containing a 100 nm focus and one a 1 mm focus, are available, each with multiple diagnostics, sample injection, pump-probe and detector capabilities. The flexibility of CXI has enabled it to host a diverse range of experiments, from biological to extreme matter.

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“…A number of room temperature sample environments tested at SR-MPX beam lines have been initially developed for SFX experiments [12,[30][31][32][33]. Such experiments are generally performed without crystal rotation.…”

Section: Riekelmentioning

confidence: 99%