title: Helicobacter pylori VacA subdomain required for intracellular toxin activity and assembly of functional oligomeric complexes.
abstract: Helicobacter pylori VacA is a secreted pore-forming toxin that is comprised of two domains, designated p33 and p55. The p55 domain has an important role in the binding of VacA to eukaryotic cell surfaces. A total of 111 residues at the amino terminus of p55 (residues 312 to 422) are essential for the intracellular activity of VacA, which suggests that this region may constitute a subdomain with an activity distinct from cell binding. To investigate the properties of this subdomain, a small deletion mutation (targeting aspartic acid 346 and glycine 347) was introduced into the H. pylori chromosomal vacA gene. Similar to wild-type VacA, the VacA Delta346-347 mutant protein was proteolytically processed, secreted, and bound to eukaryotic cells. However, VacA Delta346-347 did not cause cell vacuolation or membrane depolarization, and it was impaired in the ability to assemble into large water-soluble oligomeric structures. Interestingly, VacA Delta346-347 was able to physically interact with wild-type VacA to form mixed oligomeric complexes, and VacA Delta346-347 inhibited wild-type vacuolating activity in a dominant-negative manner. These data indicate that the assembly of functional oligomeric VacA complexes is dependent on specific sequences, including amino acids 346 and 347, within the p55 amino-terminal subdomain.
authors.affiliation: Department of Microbiology and Immunology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-2605, USA.
authors.forename: Susan E
authors.initials: SE
authors.lastname: Ivie
authors.forename: Mark S
authors.initials: MS
authors.lastname: McClain
authors.forename: Victor J
authors.initials: VJ
authors.lastname: Torres
authors.forename: Holly M Scott
authors.initials: HM
authors.lastname: Algood
authors.forename: D Borden
authors.initials: DB
authors.lastname: Lacy
authors.forename: Rong
authors.initials: R
authors.lastname: Yang
authors.forename: Steven R
authors.initials: SR
authors.lastname: Blanke
authors.forename: Timothy L
authors.initials: TL
authors.lastname: Cover
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D016480:Helicobacter pylori / Q000235:genetics / Q000254:growth & development / Q000378:metabolism* / Q000472:pathogenicity; D006801:Humans; D008564:Membrane Potentials; D009154:Mutation; D010957:Plasmids; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013329:Structure-Activity Relationship
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; C074689:VacA protein, Helicobacter pylori