Abstract
Protein crystallography using synchrotron radiation sources has had a tremendous impact on biology, having yielded the structures of thousands of proteins and given detailed insight into their mechanisms. However, the technique is limited by the requirement for macroscopic crystals, which can be difficult to obtain, as well as by the often severe radiation damage caused in diffraction experiments, in particular when using tiny crystals. To slow radiation damage, data collection is typically performed at cryogenic temperatures. With the advent of free-electron lasers (FELs) capable of delivering extremely intense femtosecond X-ray pulses, this situation appears to be remedied, allowing the structure determination of undamaged macromolecules using either macroscopic or microscopic crystals. The latter are exposed to the FEL beam in random orientations and their diffraction data are collected at cryogenic or room temperature in a serial fashion, since each crystal is destroyed upon a single exposure. The new approaches required for crystal growth and delivery, and for diffraction data analysis, including de novo phasing, are reviewed. The opportunities and challenges of SFX are described, including applications such as time-resolved measurements and the analysis of radiation damage-prone systems.
Highlights
‘It was a wonderful time’, Lawrence Bragg remembered
Radiation damage limits the amount of useful diffraction data that can be obtained from small crystals (Holton & Frankel, 2010), even when they are kept at cryogenic temperature during data collection to slow the diffusion of radiation-induced radicals
While this has remained a dream for second- and third-generation synchrotron X-ray sources, it seemed feasible for XFELs in line with molecular dynamicsbased simulations performed by Hadju and coworkers (Neutze et al, 2000)
Summary
‘It was a wonderful time’, Lawrence Bragg remembered. ‘Like discovering a new goldfield where nuggets could be picked up on the ground, with thrilling new results every week’ (Nobel prize lecture, 1915; http://www.nobelprize.org/nobel_prizes/ physics/laureates/1915/wl-bragg-lecture.html). XFELs are linear accelerator-based X-ray sources that deliver femtosecond coherent X-ray pulses with a peak brilliance that is nine orders of magnitude higher than that of third-generation synchrotron sources With these beam characteristics, the two currently available hard X-ray FELs, the Linac Coherent Light Source (LCLS) at SLAC/Stanford, USA (Emma et al, 2010), and the Spring-8 Angstrom Compact Free-electron Laser (SACLA) (Ishikawa et al, 2012) at Riken/ Harima, Japan, allow unprecedented studies in many different areas of science. Solem pointed out in 1986 (Solem, 1986) that radiation damage can be prevented if the diffraction data are acquired sufficiently rapidly While this has remained a dream for second- and third-generation synchrotron X-ray sources, it seemed feasible for XFELs in line with molecular dynamicsbased simulations performed by Hadju and coworkers (Neutze et al, 2000). Recent reviews include Fromme & Spence (2011), Patterson (2014), Schlichting & Miao (2012) and Spence et al (2012)
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