Final Activity Report Summary - RNA POLYMERASE III (Structural studies of yeast RNA polymerase III)
As stated in the original working plan, we managed to establish large-scale fermentation (200 liters) and purification of Pol III from S. cerevisiae. By means of cryo-electron microscopy and crystallography (hybrid approach) we were able to obtain three independent snapshot of Pol III: first, the apo-enzyme in isolation, second, the complex with a specific Pol III gene (elongation complex) and third, a complex with the global negative regulator Maf1. These reconstructions are of high quality and allowed us to rationalize the specific features of the Pol III enzyme and its activity, supporting previously published biochemical data, now on a structural level.
Upon completion of functional in vitro data experiments to corroborate our model, we will be able to publish our findings in a high-impact journal, given the importance of such basic mechanisms that are of central importance for the whole transcription regulation community.