Cytotoxicity of the Bacillus cereus Nhe enterotoxin requires specific binding order of its three exoprotein components.
This study focuses on the interaction of the three components of the Bacillus cereus Nhe enterotoxin with particular emphasis on the functional roles of NheB and NheC. The results demonstrated that both NheB and NheC were able to bind to Vero cells directly while NheA lacked this ability. It was also shown that Nhe-induced cytotoxicity required a specific binding order of the individual components whereby the presence of NheC in the priming step as well as the presence of NheA in the final incubation step was mandatory. Priming of cells with NheB alone and addition of NheA plus NheC in the second step failed to induce toxic effects. Furthermore, in solution, excess NheC inhibited binding of NheB to Vero cells, whereas priming of cells with excess NheC resulted in full toxicity if unbound NheC was removed before addition of NheB. By using mutated NheC proteins where the two cysteine residues in the predicted beta-tongue were replaced with glycine (NheCcys-) or where the entire hydrophobic stretch was deleted (NheChr-), the predicted hydrophobic beta-tongue of NheC was found essential for binding to cell membranes but not for interaction with NheB in solution. All data presented here are compatible with the following model. The first step in the mode of action of Nhe is associated with binding of NheC and NheB to the cell surface and probably accompanied by conformational changes. These events allow subsequent binding of NheA, leading to cell lysis.
Lindbäck
Toril
T
Department of Food Safety and Infection Biology, Norwegian School of Veterinary Science, P.O. Box 8146 Dep., N-0033 Oslo, Norway.
Hardy
Simon P
SP
Dietrich
Richard
R
Sødring
Marianne
M
Didier
Andrea
A
Moravek
Maximilian
M
Fagerlund
Annette
A
Bock
Stefanie
S
Nielsen
Carina
C
Casteel
Maximilian
M
Granum
Per Einar
PE
Märtlbauer
Erwin
E
D000818:Animals; D001409:Bacillus cereus / Q000472:pathogenicity*; D002522:Chlorocebus aethiops; D004305:Dose-Response Relationship, Drug; D004768:Enterotoxins / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D014709:Vero Cells
D004768:Enterotoxins; C040792:enterotoxin, Bacillus cereus; D007770:L-Lactate Dehydrogenase