Reproducibility of single protein explosions induced by X-ray lasers

Abstract

Single particle imaging (SPI) using X-ray pulses has become increasingly attainable with the advent of high-intensity free electron lasers. Eliminating the need for crystallized samples enables structural studies of molecules previously inaccessible by conventional crystallography. While this emerging technique already demonstrates substantial promise, some obstacles need to be overcome before SPI can reach its full potential. One such problem is determining the spatial orientation of the sample at the time of X-ray interaction. Existing solutions rely on diffraction data and are computationally demanding and sensitive to noise. In this in silico study, we explore the possibility of aiding these methods by mapping the ion distribution as the sample undergoes a Coulomb explosion following the intense ionization. By detecting the ions ejected from the fragmented sample, the orientation of the original sample should be possible to determine. Knowledge of the orientation has been shown earlier to be of substantial advantage in the reconstruction of the original structure. 150 explosions of each of twelve separate systems - four polypeptides with different amounts of surface bound water - were simulated with molecular dynamics (MD) and the average angular distribution of carbon and sulfur ions was investigated independently. The results show that the explosion maps are reproducible in both cases, supporting the idea that orientation information is preserved. Additional water seems to restrict the carbon ion trajectories further through a shielding mechanism, making the maps more distinct. For sulfurs, water has no significant impact on the trajectories, likely due to their higher mass and greater ionization cross section, indicating that they could be of particular interest. Based on these findings, we conclude that explosion data can aid spatial orientation in SPI experiments and could substantially improve the capabilities of the novel technique.

Structural Heterogeneity in Single Particle Imaging Using X-ray Lasers

“…In an earlier study we described the trajectories of the ions from a protein exploding due to the heavy ionization caused by an XFEL pulse. 13 There it was shown that the same ions from individual explosions tend to fly out in similar directions. This suggests that recording the paths of the ejected ions could be a way to determine the orientation of the protein at the moment when it is hit by the XFEL pulse.…”

“… 21 It has also been shown at the same time that encapsulating biomolecules in water will lead to a reduction of radiation damage, 22 and furthermore the Coulomb explosion of single molecules becomes more reproducible when the samples are slightly hydrated. 13 …”

Is radiation damage the limiting factor in high-resolution single particle imaging with X-ray free-electron lasers?

“…It has recently been shown that the random orientation and the low photon signal per shot can be tackled with de novo correlation approaches and that a low number of coherently scattered photons are sufficient 35 . Recent studies discuss ways to experimentally preorient a protein before the X-ray exposure, 36 or orient the sample postexposure 8 . The present study does not address the problem with finding the orientation of the sample as it is hit by the XFEL pulse but investigates the impact of damage on the diffraction patterns from aligned molecules.…”

“…Following our earlier work, 8,19,20 we simulated the interaction between a lysozyme protein molecule (PDB identifier 1LYS 21 ) and an ultrashort XFEL pulse using the molecular dynamics (MD) software GROMACS 3 22 . MD is a simulation scheme based entirely upon classical mechanics that has been shown to accurately describe such interactions 23 .…”

“…Here, we aimed to complement this by applying an equivalent methodology to simulations of an exploding molecule. Building on our recent study of the explosion dynamics of proteins exposed to an XFEL pulse, 8 we investigated how the Coulomb explosion contributes to the noise in the diffracted signal. The impact was evaluated through comparisons to noise caused by structural variations within the sample, i.e., sample heterogeneity, and due to deviations in scattering between pulses, the so-called shot noise.…”