Protein Dynamics via Time-resolved Non-linear Spectroscopy in the Ultraviolet
Ultrafast non-linear spectroscopy allows the observation of small scale structural fluctuations, which are important for our understanding of protein function and protein folding. By probing in the ultraviolet (UV), we can use the aromatic amino acids as probes, which naturally occur in proteins, thus probing directly wild-type proteins without any modification.
In this project, we want to study dynamics in myoglobin as a first system. Our initial goal is to obtain detailed information of the equilibrium dynamics in the environment of the tryptophan-residue Trp-14 in different myoglobins, to better understand the reasons for its extreme evolutionary conservation and therefore, its functional role.
In a second step, we will investigate myoglobin function by following the changes in the protein after photo-dissociation of the ligand. Probing the tryptophan in myoglobin with nonlinear optical techniques will reveal the local structural dynamics and changes due to relaxation during the protein’s functional activity.
For this goal, we developed UV photon echo techniques and applied them in combination with ultrafast time-resolved fluorescence and pump-probe techniques. Recently we have constructed a 2D UV pump-probe experiment, and performed the first 2D-UV experiments on proteins (In Press, figure below shows results obtained for a dye molecule).
The importance of extending time-resolved non-linear spectroscopic techniques to the UV can hardly be underestimated. It is not restricted to investigations on myoglobin, but has much broader applications as tryptophan is most frequently encountered in proteins. Thus, the present study will open a whole new field of investigations, applicable to any protein which contains aromatic amino acids.
J. Léonard, E. Portuondo-Campa, A. Cannizzo, F. van Mourik and G. van der Zwan et al. Functional electric field changes in photoactivated proteins revealed by ultrafast Stark spectroscopy of the Trp residues, Proceedings of the National Academy of Sciences 106, p. 7718-7723, 2009.