title: Relationship between heat-induced fibrillogenicity and hemolytic activity of thermostable direct hemolysin and a related hemolysin of Vibrio parahaemolyticus.
abstract: The formation of nonspecific ion channels by small oligomeric amyloid intermediates is toxic to the host's cellular membranes. Thermostable direct hemolysin (TDH) and TDH-related hemolysin (TRH) are major virulence factors of Vibrio parahaemolyticus. We have previously reported the crystal structure of TDH tetramer with the central channel. We have also identified the molecular mechanism underlying the paradoxical responses to heat treatment of TDH, known as the Arrhenius effect, which is the reversible amyloidogenic property. In the present report, we describe the biophysical properties of TRH, which displays 67% amino acid similarity with TDH. Molecular modeling provided a good fit of the overall structure of TDH and TRH. Size-exclusion chromatography, ultracentrifugation, and transmission electron microscopy revealed that TRH formed tetramer in solution. These toxins showed similar hemolytic activity on red blood cells. However, TRH had less amyloid-like structure than TDH analyzed by thioflavin T-binding assay and far-UV circular dichroism spectra. These data indicated that amyloidogenicity upon heating is not essential for the membrane disruption of erythrocytes, but the maintenance of tetrameric structure is indispensable for the hemolytic activity of the TDH and TRH.
authors.affiliation: Department of Developmental Medicine, Osaka Medical Center for Maternal and Child Health, Research Institute, Osaka, Japan.
authors.forename: Kiyouhisa
authors.initials: K
authors.lastname: Ohnishi
authors.forename: Kumiko
authors.initials: K
authors.lastname: Nakahira
authors.forename: Satoru
authors.initials: S
authors.lastname: Unzai
authors.forename: Kouta
authors.initials: K
authors.lastname: Mayanagi
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Hashimoto
authors.forename: Kentaro
authors.initials: K
authors.lastname: Shiraki
authors.forename: Takeshi
authors.initials: T
authors.lastname: Honda
authors.forename: Itaru
authors.initials: I
authors.lastname: Yanagihara
mesh_terms: D001427:Bacterial Toxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000633:toxicity; D002942:Circular Dichroism; D004912:Erythrocytes / Q000382:microbiology; D006460:Hemolysin Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000633:toxicity*; D006461:Hemolysis; D006358:Hot Temperature; D006801:Humans; D055503:Protein Multimerization; D055550:Protein Stability; D014736:Vibrio parahaemolyticus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*; D037521:Virulence Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000633:toxicity*
chemical_list: D001427:Bacterial Toxins; D006460:Hemolysin Proteins; D037521:Virulence Factors; C068489:thermostable direct hemolysin