Abstract
Serial femtosecond crystallography is an X-ray free-electron-laser-based method with considerable potential to have an impact on challenging problems in structural biology. Here we present X-ray diffraction data recorded from microcrystals of the Blastochloris viridis photosynthetic reaction centre to 2.8 Å resolution and determine its serial femtosecond crystallography structure to 3.5 Å resolution. Although every microcrystal is exposed to a dose of 33 MGy, no signs of X-ray-induced radiation damage are visible in this integral membrane protein structure.
Highlights
Serial femtosecond crystallography is an X-ray free-electron-laser-based method with considerable potential to have an impact on challenging problems in structural biology
The X-ray exposure for each and every microcrystal is 33 MGy, which is approximately two orders of magnitude beyond that normally suitable for room temperature data collection, no signs of radiation damage are visible in the resulting electron density. These results demonstrate the considerable potential of XFELbased SFX when applied to the challenging field of membrane protein structural biology
The lipidic sponge phase (LSP)-grown RCvir microcrystals were injected as a microjet[7,8] across the focused X-ray free-electron lasers (XFELs) beam of the Linac Coherent Light Source (LCLS) and diffraction data to 2.8 Å resolution (Fig. 1) were collected on a Cornell-SLAC Pixel Array detector[9] at the coherent X-ray imaging (CXI) instrument[16]. As this beamline operates at a short wavelength (B1.4 Å), it was possible to extend our earlier 8.2 Å resolution RCvir SFX structure[4] to 3.5 Å resolution, at which most side chains can be assigned without ambiguity
Summary
Serial femtosecond crystallography is an X-ray free-electron-laser-based method with considerable potential to have an impact on challenging problems in structural biology. Serial femtosecond crystallography (SFX) brings a new dimension to the challenge of screening small crystals This approach has been developed as a high-resolution structural method[3,4,5,6] at X-ray free-electron lasers (XFELs) and allows diffraction data to be collected at room temperature from thousands of randomly oriented, fully hydrated micron-sized crystals injected into the XFEL beam using a microjet[7,8]. High-resolution SFX structures of the soluble proteins lysozyme[5] and cathepsin B6 (1.9 and 2.1 Å resolution, respectively) were reported following the commissioning of the coherent X-ray imaging (CXI) instrument[16], which operates at shorter wavelengths Despite these advances, diffraction data recorded at the LCLS from membrane protein microcrystals have been considerably weaker than data recorded from microcrystals of soluble proteins
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