Analysis of the catalytic site of the actin ADP-ribosylating Clostridium perfringens iota toxin

The enzyme component of the actin ADP-rihosylating Clostridium perfringens iota toxin was affinity labelled by UV irradiation in the presence of [carbonyl-C-14]NAD. A peptide containing the radiolabel was generated by CNBr cleavage and subsequent proteolysis with trypsin, Its amino acid sequence is Gly-Ser-Pro-Gly-Ala-Tyr-Leu-Ser-Ala-Ile-Pro-Gly-Tyr-Ala-G-ly-X-Tyr-Glu-Val-Leu-Leu-Asn-His-Gly-Ser-Lys corresponding with the region Gly-363 through Lys-388 in the C. perfringens iota toxin. Mass spectrometric data as well as the results of the PTH-amino acid analysis are in line with a modification of a glutamic acid side chain located at position 378. Therefore, in addition to Glu-380, as could be concluded by analogy with other ADP-ribosyltransferases, Glu-378 may play a pivotal role in the active site of C. peufringens iota toxin.