title: Identification of Glu173 as the critical amino acid residue for the ADP-ribosyltransferase activity of Clostridium botulinum C3 exoenzyme.
abstract: Clostridium botulinum C3 exoenzyme specifically ADP-ribosylates rho-p21 in eukaryotic cells. Trp18 and Glu173 of this enzyme were substituted with other amino acids via site-directed mutagenesis. All substitutions at Glu173 caused a significant reduction in affinity for NAD and diminished ADP-ribosyltransferase activity. On the other hand, the activity of enzymes with the substitution at Trp18 remained intact. Swiss 3T3 cells treated with the enzyme with the Trp18 substitution showed the typical morphologic changes of the C3 exoenzyme phenotype. In contrast, no changes were found in cells incubated with the Glu173-substituted enzyme. These results indicate that the Glu173 residue of the C3 exoenzyme plays a key role in interacting with NAD and in expression of ADP-ribosyltransferase activity, which is essential for the phenotypic change by C3 exoenzyme treatment.
authors.affiliation: Department of Pharmacology, Kyoto University Faculty of Medicine, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Saito
authors.forename: Y
authors.initials: Y
authors.lastname: Nemoto
authors.forename: T
authors.initials: T
authors.lastname: Ishizaki
authors.forename: N
authors.initials: N
authors.lastname: Watanabe
authors.forename: N
authors.initials: N
authors.lastname: Morii
authors.forename: S
authors.initials: S
authors.lastname: Narumiya
mesh_terms: D016475:3T3 Cells; D036002:ADP Ribose Transferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001905:Botulinum Toxins; D018698:Glutamic Acid; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009243:NAD / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014364:Tryptophan
chemical_list: D009243:NAD; D018698:Glutamic Acid; D014364:Tryptophan; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; D011065:Poly(ADP-ribose) Polymerases; D001905:Botulinum Toxins