The N-terminus of the Clostridioides difficile transferase A component directs toxin activity and potency

Clostridioides difficile infection is the leading cause of antibiotic-associated, hospital-acquired diarrhea in the USA; the pathology of which is mediated by toxins. The presence of a toxin known as the C. difficile Transferase (CDT) in some clinical isolates is linked to severe symptoms including increased incidence of reinfection and higher rates of mortality. Despite its apparent importance to C. difficile pathology, a mechanistic model of how CDT intoxicates cells remains incomplete. Here, we describe a motif composed of acidic and basic residues (the KDKEK motif) that is essential for toxin function. Using Cryogenic Electron Microscopy (Cryo-EM), we highlight an orientation of the KDKEK motif wherein the acidic residues engage structures thought to play an important role during toxin delivery. We thus present a model wherein these interactions prime CDT for entry into host cells. We expect that this model can be extrapolated to other bacterial toxins to understand how they enter cells.