title: Crystal structure of the C3bot-RalA complex reveals a novel type of action of a bacterial exoenzyme.
abstract: C3 exoenzymes from bacterial pathogens ADP-ribosylate and inactivate low-molecular-mass GTPases of the Rho subfamily. Ral, a Ras subfamily GTPase, binds the C3 exoenzymes from Clostridium botulinum and C. limosum with high affinity without being a substrate for ADP ribosylation. In the complex, the ADP-ribosyltransferase activity of C3 is blocked, while binding of NAD and NAD-glycohydrolase activity remain. Here we report the crystal structure of C3 from C. botulinum in a complex with GDP-bound RalA at 1.8 A resolution. C3 binds RalA with a helix-loop-helix motif that is adjacent to the active site. A quaternary complex with NAD suggests a mode for ADP-ribosyltransferase inhibition. Interaction of C3 with RalA occurs at a unique interface formed by the switch-II region, helix alpha3 and the P loop of the GTPase. C3-binding stabilizes the GDP-bound conformation of RalA and blocks nucleotide release. Our data indicate that C. botulinum exoenzyme C3 is a single-domain toxin with bifunctional properties targeting Rho GTPases by ADP ribosylation and Ral by a guanine nucleotide dissociation inhibitor-like effect, which blocks nucleotide exchange.
authors.affiliation: Structural Research, Department of Integrated Lead Discovery, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riss, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Pautsch
authors.forename: Martin
authors.initials: M
authors.lastname: Vogelsgesang
authors.forename: Jens
authors.initials: J
authors.lastname: Tränkle
authors.forename: Christian
authors.initials: C
authors.lastname: Herrmann
authors.forename: Klaus
authors.initials: K
authors.lastname: Aktories
mesh_terms: D036002:ADP Ribose Transferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000227:Adenine Nucleotides / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001905:Botulinum Toxins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003014:Clostridium botulinum / Q000378:metabolism*; D003461:Crystallography; D019281:Dimerization; D008969:Molecular Sequence Data; D009154:Mutation; D009243:NAD / Q000378:metabolism; D011487:Protein Conformation; D020725:ral GTP-Binding Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: D000227:Adenine Nucleotides; D009243:NAD; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; D001905:Botulinum Toxins; D020725:ral GTP-Binding Proteins