Cryo-EM structural studies of OdhA, a large, multifunctional actinobacterial 2-oxoglutarate dehydrogenase

The purpose of this proposal is to characterize the OdhA enzyme (2-oxoglutarate dehydrogenase) from the model actinobacterium Corynebacterium glutamicum. This peculiar enzyme has several features: it is a fusion enzyme carrying two distinct activities (normally carried out by distinct enzymes), it possesses a peculiar domain organization, and it is regulated through a signal transduction cascade that is also proper to actinobacteria. In addition, OdhA is part of a larger, still uncharacterized PDH/ODH mixed supercomplex which represents another actinobacterial feature. We specifically propose here to make use of the ESRF CM01 facility to collect 300kV data on optimized grids of both full-length OdhA with substrate analogues, and the OdhA-OdhI complex. We aim at reaching nearly atomic resolution for both complexes, allowing to unravel the mechanistic aspects of the peculiar regulation of this large enzymatic machinery.