The central project in our laboratory is currently the investigation of the molecular machine, which releases many neurotransmitters and hormones in the body. We investigate the dynamics of vesicle docking, priming and fusion dependent on SNAREs and accessory proteins in live cells using electrophysiological, electrochemical and advanced fluorescence imaging techniques as well as molecular dynamics simulations. Our approaches include microfabrication of electrochemical detector (ECD) arrays and their use in combination with Total Internal Reflection (TIRF) microscopy.
One approach uses whole cell patch clamp capacitance measurements to investigate the functional performance of fluorescently labeled proteins in knock-out mouse embryonal chromaffin cells and the function of otherwise modified proteins involved in vesicle priming and fusion.
A second approach combines amperometry using ECD arrays TIRF imaging to study the molecular mechanism of vesicle fusion and transmitter release in chromaffin cells.
A third approach combines ECD arrays, with reconstituted supported membranes to study fusion of isolated chromaffin granules. This reconstituted system will make it possible to incorporate small labels at arbitrary sites in the SNARE proteins or other accessory proteins, to identify precisely which amino acids in the SNARE complex and accessory proteins move and change distance at specific times during the fusion process.
In a 4th approach we perform Molecular Dynamics modeling of the SNARE complex and accessory proteins to elucidate the changes in protein interactions and conformations leading to vesicle fusion.
