title: Enzymatic actions of Pasteurella multocida toxin detected by monoclonal antibodies recognizing the deamidated α subunit of the heterotrimeric GTPase Gq.
abstract: Pasteurella multocida toxin (PMT) is a virulence factor responsible for the pathogenesis of some Pasteurellosis. PMT exerts its toxic effects through the activation of heterotrimeric GTPase (G(q), G(12/13) and G(i))-dependent pathways, by deamidating a glutamine residue in the α subunit of these GTPases. However, the enzymatic characteristics of PMT are yet to be analyzed in detail because the deamidation has only been observed in cell-based assays. In the present study, we developed rat monoclonal antibodies, specifically recognizing the deamidated Gα(q), to detect the actions of PMT by immunological techniques such as western blotting. Using the monoclonal antibodies, we found that the toxin deamidated Gα(q) only under reducing conditions. The C-terminal region of PMT, C-PMT, was more active than the full-length PMT. The C3 domain possessing the enzyme core catalyzed the deamidation in vitro without any other domains. These results not only support previous observations on toxicity, but also provide insights into the enzymatic nature of PMT. In addition, we present several lines of evidence that Gα(11), as well as Gα(q), could be a substrate for PMT.
authors.affiliation: Department of Molecular Bacteriology, Research Institute for Microbial Diseases, Osaka University, Suita-shi, Osaka, Japan. skami@biken.osaka-u.ac.jp
authors.forename: Shigeki
authors.initials: S
authors.lastname: Kamitani
authors.forename: Shinpei
authors.initials: S
authors.lastname: Ao
authors.forename: Hirono
authors.initials: H
authors.lastname: Toshima
authors.forename: Taro
authors.initials: T
authors.lastname: Tachibana
authors.forename: Makiko
authors.initials: M
authors.lastname: Hashimoto
authors.forename: Kengo
authors.initials: K
authors.lastname: Kitadokoro
authors.forename: Aya
authors.initials: A
authors.lastname: Fukui-Miyazaki
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Abe
authors.forename: Yasuhiko
authors.initials: Y
authors.lastname: Horiguchi
mesh_terms: D016475:3T3 Cells; D000818:Animals; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins / Q000276:immunology / Q000378:metabolism*; D001427:Bacterial Toxins / Q000276:immunology / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D051381:Rats
chemical_list: D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D001427:Bacterial Toxins; C062258:Pasteurella multocida toxin; D043802:GTP-Binding Protein alpha Subunits, Gq-G11