Tc toxins deliver toxic enzymes into host cells by a unique injection mechanism. One of these enzymes is the actin ADP-ribosyltransferase TccC3, whose activity leads to the clustering of the cellular cytoskeleton and ultimately cell death. Here, we show in atomic detail how TccC3 modifies actin. Here the Raunser lab finds that the ADP-ribosyltransferase does not bind to G-actin but interacts with two consecutive actin subunits of F-actin. The binding of TccC3 to F-actin occurs via an induced-fit mechanism that facilitates access of NAD+ to the nucleotide binding pocket. The following nucleophilic substitution reaction results in the transfer of ADP-ribose to threonine-148 of F-actin. We demonstrate that this site-specific modification of F-actin prevents its interaction with depolymerization factors, such as cofilin, which impairs actin network turnover and leads to steady actin polymerization. Our findings reveal in atomic detail a mechanism of action of a bacterial toxin through specific targeting and modification of F-actin.
After binding of the toxin to its receptor(s) on the surface of the target cell, the Tc complex is endocytosed. Inside the late endosome, the Tc toxin undergoes the prepore-to-pore transition and the TcHVR is translocated into the host cell cytoplasm. There, TcHVR folds and interacts with F-actin via an induced-fit mechanism that facilitates NAD+ access to the nucleotide-binding pocket. The following reaction results in the transfer of ADP-ribose to threonine-148 of F-actin. The modification impairs the interaction of F-actin with actin-depolymerizing factors, which inhibits actin turnover and eventually leads to cell death.
After binding of the toxin to its receptor(s) on the surface of the target cell, the Tc complex is endocytosed. Inside the late endosome, the Tc toxin undergoes the prepore-to-pore transition and the TcHVR is translocated into the host cell cytoplasm. There, TcHVR folds and interacts with F-actin via an induced-fit mechanism that facilitates NAD+ access to the nucleotide-binding pocket. The following reaction results in the transfer of ADP-ribose to threonine-148 of F-actin. The modification impairs the interaction of F-actin with actin-depolymerizing factors, which inhibits actin turnover and eventually leads to cell death.