Identification of the C-terminal part of Bordetella dermonecrotic toxin as a transglutaminase for rho GTPases

Bordetella dermonecrotic toxin (DNT) causes the deamidation of glutamine 63 of Rho. Here we identified the region of DNT harboring the enzyme activity and compared the toxin with the cytotoxic necrotizing factor 1, which also deamidates Rho. The DNT fragment (Delta DNT) covering amino acid residues 1136-1451 caused deamidation of RhoA at glutamine 63 as determined by mass spectrometric analysis and by the release of ammonia. In the presence of dansylcadaverine or ethylenediamine, Delta DNT caused transglutamination of Rho. Deamidase and transglutaminase activities were blocked in the mutant proteins Cys(1292) --> Ala, His(1307) --> Ala, and Lys(1310) --> Ala of Delta DNT. Deamidation and transglutamination induced by Delta DNT blocked intrinsic and Rho-GTPase-activating protein-stimulated GTPase activity of RhoA. Delta DNT deamidated and transglutaminated Rac and Cdc42 in the absence and presence of ethylenediamine, respectively. Modification of Rho proteins by Delta DNT was nucleotide-dependent and did not occur with GTP gamma S-loaded GTPases, In contrast to cytotoxic necrotizing factor, which caused the same kinetics of ammonia release in the absence and presence of ethylenediamine, ammonia release by Delta DNT was largely increased in the presence of ethylenediamine, indicating that Delta DNT acts primarily as a transglutaminase.