Research group Tilman Schirmer
Molecular mechanisms of c-di-GMP signal transduction and AMP transferases
We are employing crystallographic and biochemical/ biophysical techniques to reveal the structural basis for the catalysis and regulation of c-di-GMP related proteins. Our second focus is on bacterial type IV secretion system (T4SS) effector proteins with AMP transferase activity.
Diguanylate cyclases and regulation of c-di-GMP synthesis
Recent discoveries show that a novel second messenger, c-di-GMP, is extensively used by bacteria to control multicellular behavior, such as biofilm formation. Condensation of two GTP to the dinucleotide is catalyzed by GGDEF domains that usually occur in combination with sensory and/or regulatory modules. The opposing phosphodiesterase activity is provided by EAL domains that are also regulated.
In collaboration with the Jenal group (Biozentrum) and based on crystallographic and functional studies we have studied the catalytic and regulatory mechanisms of diguanylate cyclades, the enzymes that synthesize the second messenger. It appears that the general mechanism of activation relies on signal induced dimerisation of its regulatory domains that ensures productive of the two GTP loaded catalytic GGDEF domains. Though the substrate binding site of c-di-GMP specific phosphodiesterases is completely contained with the EAL domain, the domain is active only as a homodimer. This generic property of the catalytic domain is probably utilised in the full-length proteins to control their activity,very similar to the situation in diguanylate cyclades.
Thus our results provide clues about how this class of enzymes can be regulated in a modular and universal fashion by their sensory domains.
Recently we have started to elucidate the structures and binding modes of newly discovered c-di-GMP receptors. These results will add to our knowledge of complete c-di-GMP signal cascades.
Effector proteins of the type IV secretion system
Type IV secretion systems (T4SS) are utilized by many bacterial pathogens for the delivery of virulence proteins or protein-DNA complexes into their eukaryotic target cells. Together with the Dehio group (Biozentrum) we are working on a class of effector proteins that are composed of a Fic and a BID domain responsible for pathogenic action in the host cell and translocation, respectively.
Only recently, it has become apparent that the Fic domain catalyzes AMP transfer onto host target protein(s) to subvert cellular function. From a Fic crystal structure (truncated BepA from Bartonella henselae) we were able to deduce the mechanisms of catalysis and target positioning. Currently, we are investigating Fic inhibition that - depending on the protein - is caused by an α-helix that interferes with productive binding of the ATP substrate or, inter-molecularly, by complex formation with an anti-toxin. Interestingly, both inhibition mechanisms are structurally related. This knowledge may be utilized for drug development to target Fic proteins of bacterial pathogens.
Porins are integral membrane proteins from the outer membrane of Gram-negative bacteria. They allow the uptake of nutrients by passive diffusion through an intrinsic pore that extends along the axis of the transmembrane β-barrel structure. After extensive work on the general trimeric porins OmpF and OmpC from E. coli, we have recently determined the high-resolution 12-stranded β-barrel structures of NanC from E. coli and KdgM from Dickeya dadantii, representatives of a porin family that is specific for the translocation of negatively charged poly-saccharides.