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 (DeltaDNT) 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, DeltaDNT 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 DeltaDNT. Deamidation and transglutamination induced by DeltaDNT blocked intrinsic and Rho- GTPase-activating protein-stimulated GTPase activity of RhoA. DeltaDNT deamidated and transglutaminated Rac and Cdc42 in the absence and presence of ethylenediamine, respectively. Modification of Rho proteins by DeltaDNT was nucleotide-dependent and did not occur with GTPgammaS-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 DeltaDNT was largely increased in the presence of ethylenediamine, indicating that DeltaDNT acts primarily as a transglutaminase.
Schmidt
G
G
Institut für Pharmakologie und Toxikologie der Albert-Ludwigs-Universität Freiburg, Hermann-Herder-Strasse 5, D-79104 Freiburg, Germany.
Goehring
U M
UM
Schirmer
J
J
Lerm
M
M
Aktories
K
K
D000595:Amino Acid Sequence; D001427:Bacterial Toxins / Q000378:metabolism*; D001665:Binding Sites; D001884:Bordetella / Q000201:enzymology*; D005029:Ethylenediamines / Q000378:metabolism; D005973:Glutamine / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013329:Structure-Activity Relationship; D011503:Transglutaminases / Q000378:metabolism*; D010566:Virulence Factors, Bordetella; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000378:metabolism*
D001427:Bacterial Toxins; D005029:Ethylenediamines; D010446:Peptide Fragments; D010566:Virulence Factors, Bordetella; C060969:dermonecrotic toxin, Bordetella; D005973:Glutamine; C031234:ethylenediamine; D011503:Transglutaminases; D020764:cdc42 GTP-Binding Protein; D020741:rho GTP-Binding Proteins