title: Formation of small transmembrane pores: An intermediate stage on the way to Bacillus cereus non-hemolytic enterotoxin (Nhe) full pores in the absence of NheA.
abstract: The non-hemolytic enterotoxin (Nhe) of Bacillus cereus is a three-partite toxin formed of the components NheA, -B and -C. Pore formation and subsequent lysis of target cells caused by Nhe is an orchestrated process comprising three steps: (i) formation of NheB/C oligomers in solution, (ii) attachment of the oligomers to the cell membrane, (iii) binding of NheA to the oligomers. The present study aimed to characterize the properties of the NheB/C complex and the fate of the target cell upon binding. An enzyme immunoassay allowing kinetic measurements and surface plasmon resonance revealed the fast and high affinity formation of the NheB/C oligomers. The benefit of these complexes is a more stable cell binding as well as stronger and earlier cytotoxic effect. High molecular mass hetero-oligomers (620 kDa) probably consisting of one NheC and up to 15 NheB were detected by size-exclusion chromatography and on native PAGE immunoblots. Due to the NheBC application the morphology and membrane permeability of Vero cells is partly disturbed. Formation of stable transmembrane channels with a conductance of about 870 pS and a diameter of about 2 nm due to the application of NheBC could be demonstrated in lipid bilayer experiments. Thus, the NheBC complex itself has a tendency to increase the membrane permeability prior to the emergence of full pores containing also NheA.
authors.affiliation: Institute of Food Safety, Department of Veterinary Sciences, Ludwig-Maximilians-University Munich, 85764 Oberschleißheim, Germany.
authors.forename: Kui
authors.initials: K
authors.lastname: Zhu
authors.affiliation: Institute of Food Safety, Department of Veterinary Sciences, Ludwig-Maximilians-University Munich, 85764 Oberschleißheim, Germany. Electronic address: a.didier@mh.vetmed.uni-muenchen.de.
authors.forename: Andrea
authors.initials: A
authors.lastname: Didier
authors.affiliation: Institute of Food Safety, Department of Veterinary Sciences, Ludwig-Maximilians-University Munich, 85764 Oberschleißheim, Germany.
authors.forename: Richard
authors.initials: R
authors.lastname: Dietrich
authors.affiliation: Institute of Food Safety, Department of Veterinary Sciences, Ludwig-Maximilians-University Munich, 85764 Oberschleißheim, Germany.
authors.forename: Uta
authors.initials: U
authors.lastname: Heilkenbrinker
authors.affiliation: Department of Life Sciences and Chemistry, Jacobs-University Bremen, 28759 Bremen, Germany.
authors.forename: Eva
authors.initials: E
authors.lastname: Waltenberger
authors.affiliation: Institute of Food Safety, Department of Veterinary Sciences, Ludwig-Maximilians-University Munich, 85764 Oberschleißheim, Germany.
authors.forename: Nadja
authors.initials: N
authors.lastname: Jessberger
authors.affiliation: Institute of Food Safety, Department of Veterinary Sciences, Ludwig-Maximilians-University Munich, 85764 Oberschleißheim, Germany.
authors.forename: Erwin
authors.initials: E
authors.lastname: Märtlbauer
authors.affiliation: Department of Life Sciences and Chemistry, Jacobs-University Bremen, 28759 Bremen, Germany.
authors.forename: Roland
authors.initials: R
authors.lastname: Benz
mesh_terms: D000818:Animals; D001409:Bacillus cereus / Q000502:physiology*; D001426:Bacterial Proteins / Q000378:metabolism*; D001427:Bacterial Toxins / Q000378:metabolism*; D002463:Cell Membrane Permeability / Q000502:physiology*; D002522:Chlorocebus aethiops; D004768:Enterotoxins / Q000378:metabolism*; D008560:Membrane Fluidity / Q000502:physiology*; D014709:Vero Cells
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004768:Enterotoxins; C000589183:NheA protein, Bacillus cereus; C040792:enterotoxin, Bacillus cereus
keywords: Bacillus cereus; Cytotoxicity; Lipid-bilayer membrane; Non-hemolytic enterotoxin; Pore-forming toxins