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Review
. 2012 Oct;22(5):613-26.
doi: 10.1016/j.sbi.2012.07.015. Epub 2012 Aug 22.

Emerging opportunities in structural biology with X-ray free-electron lasers

Ilme Schlichting  1 , Jianwei Miao
Affiliations
Review

Emerging opportunities in structural biology with X-ray free-electron lasers

Ilme Schlichting et al. Curr Opin Struct Biol. 2012 Oct.

Abstract

X-ray free-electron lasers (X-FELs) produce X-ray pulses with extremely brilliant peak intensity and ultrashort pulse duration. It has been proposed that radiation damage can be 'outrun' by using an ultra intense and short X-FEL pulse that passes a biological sample before the onset of significant radiation damage. The concept of 'diffraction-before-destruction' has been demonstrated recently at the Linac Coherent Light Source, the first operational hard X-ray FEL, for protein nanocrystals and giant virus particles. The continuous diffraction patterns from single particles allow solving the classical 'phase problem' by the oversampling method with iterative algorithms. If enough data are collected from many identical copies of a (biological) particle, its three-dimensional structure can be reconstructed. We review the current status and future prospects of serial femtosecond crystallography (SFX) and single-particle coherent diffraction imaging (CDI) with X-FELs.

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Figures

Figure 1
Figure 1
(a) Experimental setup for serial femtosecond crystallography. Tiny crystals are injected into an X-FEL beam using a liquid microjet. This setup is very convenient for pump-probe experiments using laser excitation. Shown is an experiment on the photo-induced dissociation of photosystem I-ferredoxin cocrystals, taking place on a microsecond time-scale (from ref [43]). (b) Microcrystalline sample for serial femtosecond crystallography data collection. Lysozyme crystals (≤1 × 1 × 3 μm3) as seen through a conventional light microscope, (c) Microcrystalline lysozyme suspended in solution (left) and after settling (right) (from ref.[42]). It is important to note that the quantities depicted here are by no means excessive; several milliliters of very dense crystalline slurry are currently required for SFX experiments.
Figure 2
Figure 2
Simulated diffraction patterns of photosystem I (a). (b) The molecular transform of the molecule is continuous. The interference of several molecules arranged in a lattice of 2×2×2 (c), 4×4×4 (c) and 8×8×8 (e) results in the appearance of Bragg peaks and the fringes between the Bragg peaks.
Figure 3
Figure 3
Single particle coherent diffraction imaging of herpesvirus virions. (a) Coherent X-ray diffraction pattern acquired from a single, herpesvirus unstained virion. (b) The diffraction pattern was directly phased to obtain an image with a resolution of ~22 nm. (c) SEM image of the same virion. (d) Quantitative characterization of the reconstructed electron density map of the herpesvirus virion. [From ref. 82]
Figure 4
Figure 4
(a) Schematic layout of aerosolized particles injected into the X-FEL beam in random orientations. The injected particles are intercepted by the X-FEL pulses and the diffractions patterns are measured by a set of pnCCD detectors. Coherent X-ray diffraction patterns of a large aggregate (b) a water droplet (c), single T4 phage particles (d, e), a nanorice grain (f), and two nanogrrains (g), measured with the LCLS pulses. (h-j) Three representative coherent X-ray diffraction patterns measured from single mimivirus particles using LCLS pulses. The diffraction patterns were directly phased to obtain images with a resolution of ~32 nm (insets). [From refs. 84 and 86]
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