Abstract
Serial femtosecond X-ray crystallography (SFX) has created new opportunities in the field of structural analysis of protein nanocrystals. The intensity and timescale characteristics of the X-ray free-electron laser sources used in SFX experiments necessitate the analysis of a large collection of individual crystals of variable shape and quality to ultimately solve a single, average crystal structure. Ensembles of crystals are commonly encountered in powder diffraction, but serial crystallography is different because each crystal is measured individually and can be oriented via indexing and merged into a three-dimensional data set, as is done for conventional crystallography data. In this way, serial femtosecond crystallography data lie in between conventional crystallography data and powder diffraction data, sharing features of both. The extremely small sizes of nanocrystals, as well as the possible imperfections of their crystallite structure, significantly affect the diffraction pattern and raise the question of how best to extract accurate structure-factor moduli from serial crystallography data. Here it is demonstrated that whole-pattern fitting techniques established for one-dimensional powder diffraction analysis can be feasibly extended to higher dimensions for the analysis of merged SFX diffraction data. It is shown that for very small crystals, whole-pattern fitting methods are more accurate than Monte Carlo integration methods that are currently used.
Highlights
Serial femtosecond X-ray crystallography (SFX) (Chapman et al, 2011; Spence et al, 2012), which emerged with the commissioning of hard X-ray free-electron laser (XFEL) sources, provides a unique opportunity for modern biology to conduct structural analysis of proteins which have previously been inaccessible to study because of the extremely small size of crystals that they form
SFX experiments are performed in this manner due to the destructive nature of the XFEL source for which a single exposure can be expected to cause the disintegration of a nanocrystal (Neutze et al, 2000)
Presented here is an approach that builds upon an established analysis technique in powder diffraction, the whole-pattern fitting method (Le Bail et al, 1988)
Summary
Serial femtosecond X-ray crystallography (SFX) (Chapman et al, 2011; Spence et al, 2012), which emerged with the commissioning of hard X-ray free-electron laser (XFEL) sources, provides a unique opportunity for modern biology to conduct structural analysis of proteins which have previously been inaccessible to study because of the extremely small size of crystals that they form (e.g. submicron). The SFX technique involves illuminating a stream of randomly oriented protein crystals of various sizes and orientations by an extremely bright and ultra-short (tens or hundreds of femtoseconds) XFEL source and properly merging the obtained diffraction data. Whilst some characteristics of single crystals can be obtained from processing individual SFX diffraction patterns, the solution of a three-dimensional crystal structure requires the processing of large numbers of SFX diffraction patterns. The solution obtained is an average crystal unitcell structure found from data that are intrinsically based on distributions of both crystal sizes and qualities
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