Structure of the Rho-activating domain of Escherichia coli cytotoxic necrotizing factor 1

Certain uropathogenic and neonatal meningitis-causing strains of Escherichi a coli express a 114 kDa protein toxin called cytotoxic necrotizing factor 1 (CNF1). The toxin causes alteration of the host cell actin cytoskeleton a nd promotes bacterial invasion of blood-brain barrier endothelial cells. CN F1 belongs to a unique group of large cytotoxins that cause constitutive ac tivation of Rho guanosine triphosphatases (GTPases), which are key regulato rs of the actin cytoskeleton. This group also includes E. coli cytotoxic ne crotizing factor 2 (CNF2, 114 kDa) and dermonecrotic toxins (DNT, 159 kDa) of Bordetella spp. with related sequences occuring in Yersinia spp. Here we show that the catalytic region of CNF1 exhibits a novel protein fold as de termined by its 1.83 Angstrom resolution crystal structure. The structure r eveals that CNF1 has a Cys-His-main chain oxygen catalytic triad reminiscen t of enzymes belonging to the catalytic triad superfamily. The position of the catalytic Cys residue at the base of a deep pocket restricts access to potential substrates and helps explain the high specificity of this and rel ated toxins.