Reproducibility of protein explosion imaging using X-ray lasers

Sammanfattning

Determining the structure of proteins is important for understanding their function and advancing drug development. Today, X-ray Free Electron Lasers (XFELs) enable imaging of proteins through Single Particle Imaging (SPI), where ultrashort X-ray pulses produce diffraction patterns that carry information about the original structure. However, the same high-intensity radiation that enables imaging also causes rapid ionisation of the protein, leading to a Coulomb explosion where the positively charged ions repel each other and the structure rapidly fragments. Recent work has shown that these explosions can encode structural information, and are reproducible under the same beam conditions. This project aims to evaluate how varying the X-ray pulse, as well as structural heterogeneity of proteins, affect reproducibility. Reproducibility was quantified using two metrics: angular root mean square deviation (RMSD), which measures directional variability of ion trajectories, and detector pattern RMSD, which quantifies explosion map variability. Molecular dynamics simulations using MolDStruct were performed on the MS2 bacteriophage coat protein across intensities of 1010-1013 photons, with constant pulse duration at 10 fs as well as variable pulse durations of 10-100 fs with constant intensity at 1011 photons. Two datasets were analysed, a homogeneous dataset using a single fixed structure and a heterogeneous dataset with 50 structures sampled from an equilibrium simulation. The results showed that the RMSD decreased with increasing intensity, saturating when structural heterogeneity became the dominant factor of variability. Shorter pulse durations gave lower RMSD. Furthermore, the correlation observed between the angular RMSD and the detector RMSD indicates that the variability of the ion trajectories is reflected in the observable detector patterns.