The project led to the following results:
In protein diffusion:
o Used Megahertz X-ray Photon Correlation Spectroscopy at EuXFEL to measure ferritin collective diffusion at nanometer length scales and microsecond delay times, approaching the protein interaction time.
o Observed anomalous diffusion at elevated concentrations
o Proved signatures of a crossover between short-time and long-time diffusion regimes, consistent with transient cage effects arising from intermolecular constraints in highly crowded environments.
In protein crystallization:
o We proved that ferritin crystallizes in the presence of polyethylene glycol (PEG) at a wide range of molecular weights.
o A state diagram of the PEG-ferritin system has been established.
o Melting of protein crystals has been seen for large PEG molecular weight (>1000 g/mol)
o Two different mechanism of ferritin crystallization have been observed:
• For PEG 3300 and larger: consistent with classical nucleation theory or two step mechanisms
• For PEG 1000 consistent with slow ordering or two step mechanism
The work performed at European XFEL opens the possibility to measure other biological solutions in crowded state at protein molecular length scales (in the range of nanometers) and microseconds time scales. This fills the experimental gap that was present when accessing the collective diffusion in the microsecond region. This has an impact for crowded protein solutions, with possible applications in molecular biology, materials science, food and pharmaceutical industries.
In the context of crystallization, the knowledge gained with using the acoustic levitator, the ferritin-PEG system, and measuring diffusion of protein with X-ray Free Electron Lasers is going to be combine to perform an experiment in which the diffusion of the proteins in monitored during crystallization in the levitated droplets.
