title: Clostridium perfringens TpeL glycosylates the Rac and Ras subfamily proteins.
abstract: Clostridium perfringens TpeL belongs to a family of large clostridial cytotoxins that encompasses Clostridium difficile toxin A (TcdA) and B (TcdB) and Clostridium sordellii lethal toxin (TcsL). We report here the identification of the TpeL-catalyzed modification of small GTPases. A recombinant protein (TpeL1-525) derived from the TpeL N-terminal catalytic domain in the presence of streptolysin O (SLO) induced the rounding of Vero cells and the glycosylation of cellular Rac1. Among several hexoses tested, UDP-N-acetyl-glucosamine (UDP-GlcNAc) and UDP-glucose (UDP-Glc) served as cosubstrates for TpeL1-525-catalyzed modifications. TpeL1-525 catalyzed the incorporation of UDP-Glc into Ha-Ras, Rap1B, and RalA and of UDP-GlcNAc into Rac1, Ha-Ras, Rap1B, and RalA. In Rac1, TpeL and TcdB share the same acceptor amino acid for glycosylation, Thr-35. In Vero cells treated with TpeL1-525 in the presence of SLO, glycosylation leads to a translocation of the majority of Rac1 and Ha-Ras to the membrane. We demonstrate for first time that TpeL uses both UDP-GlcNAc and UDP-Glc as donor cosubstrates and modifies the Rac1 and Ras subfamily by glycosylation to mediate its cytotoxic effects.
authors.affiliation: Department of Microbiology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro-cho, Tokushima 770-8514, Japan. nagahama@ph.bunri-u.ac.jp
authors.forename: Masahiro
authors.initials: M
authors.lastname: Nagahama
authors.forename: Akiko
authors.initials: A
authors.lastname: Ohkubo
authors.forename: Masataka
authors.initials: M
authors.lastname: Oda
authors.forename: Keiko
authors.initials: K
authors.lastname: Kobayashi
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Amimoto
authors.forename: Kazuaki
authors.initials: K
authors.lastname: Miyamoto
authors.forename: Jun
authors.initials: J
authors.lastname: Sakurai
mesh_terms: D000818:Animals; D001427:Bacterial Toxins / Q000378:metabolism*; D002522:Chlorocebus aethiops; D003016:Clostridium perfringens / Q000235:genetics / Q000378:metabolism*; D003603:Cytotoxins / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000378:metabolism; D006031:Glycosylation; D011485:Protein Binding; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D011994:Recombinant Proteins; D013379:Substrate Specificity; D014709:Vero Cells; D018631:ras Proteins / Q000378:metabolism*
chemical_list: D001427:Bacterial Toxins; D003603:Cytotoxins; D011994:Recombinant Proteins; D051057:Proto-Oncogene Proteins c-akt; D020558:GTP Phosphohydrolases; D018631:ras Proteins