Research

Domain function analysis of clostridial glucosyltransferases

Our research group primarily investigates the large clostridial glucosyltransferases, which are considered to be the main pathogenicity factors of clinically important clostridia. These are toxins A and B, which cause the disease-related symptoms of C. difficile-associated diarrhea, but also the lethal and hemorrhagic toxin of C. sordellii, which cause necrosis and edema in gangrene. All clostridial glucosyltransferases modify intracellular signaling proteins of the Ras and Rho proteins, which belong to the large group of low molecular weight monomeric GTP-binding proteins. In order to reach the substrate proteins in the cytosol of the target cell, the toxins have sophisticated mechanisms for introducing their pathogenic domain into the cytosol. This includes the partially redundant binding to different receptors, the pH-dependent translocation across the endosomal membrane and the stimulable autoproteolytic cleavage of the enzymatically active domain. These tasks are performed by at least four structurally distinct domains in the toxins.

We manipulate the specific functions of the toxins by targeted genetic modifications, such as the introduction of point mutations, the exchange of domains of homologous toxins or the production of toxin chimeras. The modification of receptor specificity, autoproteolysis or substrate-modifying properties helps to understand the significance of the functional domains for the pathogenic effect of the toxins. In addition, it can be deduced why different toxin types from clinical isolates differ in their pathogenic effect with regard to the inflammatory process and the severity of the diseases.

Our current third-party funded projects focus on the interaction of the receptor binding domains with the specific cell surface receptors as well as on the intracellular localization and residence time of the pathogenic glucosyltransferase domain. Of particular interest are the effects of the toxins, which they exert independently of the glucosylation of the Rho and Ras GTPases. The endosome-associated effect on cell viability is in the foreground.

It is not only the large clostridial glucosyltransferases that have a specific uptake mechanism. All intracellularly active protein toxins have a specialized receptor binding and translocation domain, which can also be present as an independent protein, as is the case with binary toxins. The findings from our research can be used to create cell-specific tools from the toxins that are capable of transporting peptides or proteins into the cells. Our research group is also interested in using both the glucosyltransferases and the binary toxin CDT of C. difficile as molecular tools.