Electromagnetic Field Induced Out-of-Equilibrium Structural Dynamics in Protein Crystals: From Picoseconds to Milliseconds

Abstract

Light-induced interactions in biomolecules are strongly varying over different model systems. Most amino acids and proteins absorb ultraviolet light, but only a few with specialised chromophores or fluorophores, are able to absorb light in the visible spectrum. In the terahertz frequency spectrum, spectroscopy have revealed protein-light interactions which are attributed to low-frequency protein vibrations. Despite the fact that questions regarding a protein’s structure and function are integrally connected, only a small fraction have been answered. With the develop ment of ultra-fast facilities, the possibility to answer these questions at picosecond and sub-picosecond time scales are possible. The aim of this thesis is to demonstrate results from different aspects of the structural dynamics research field. This thesis show induced structural dynamics in the model protein bovine trypsin, while irradiated by a terahertz electromagnetic field at millisecond and at femtosecond timescales, during two different X-ray crystallography experiments. At millisecond timescales, differences in averages over the model parameters B-factors and anisotropy (’ANISO’) reveal structural dynamics which are not attributed to thermal vibrations. A clustering of the individual components of the anisotropic displacement parameter tensor, group atoms which have seemingly no spatial correlation. This indicates long range vibrations, oscillating over the entire protein scaffold. At femtosecond timescales, averages of distances of individual atom positions and individual B-factors, show structural differences which are distributed over the entire protein model, but localised to specific residues, or nearest neighbours. In addition, the structural dynamics in photosyntetic reaction center protein were demonstrated in a femtosecond optical pump-X-ray probe experiment. Average distance ratios of individual Cα atoms from photo-activated and ”dark” datasets were compared. The distances reveal a structural difference upon excitation with infrared light, attributed to an electron charge transfer from the special pair of chlorophyll molecules, to the menaquinone, via the cofactors of the L-subunit. The structural differences are supported by electron difference maps, time-resolved infrared spectroscopy and molecular modelling. Finally, this thesis demonstrate X-ray data, collected at a commissioning beamtime at the FemtoMAX beamline, of the short-pulse facility at MAX IV. Despite technical and practical difficulties, high resolution X-ray diffraction data were collected with good data reduction and refinement statistics. From this data, a satisfactory protein model was obtained