X-ray crystal structure of the B component of Hemolysin BL fromBacillus cereus

Biology Department, Brookhaven National Laboratory, Upton, New York 11973, USA.
Proteins Structure Function and Bioinformatics (Impact Factor: 2.63). 05/2008; 71(2):534-40. DOI: 10.1002/prot.21888
Source: PubMed


Bacillus cereus Hemolysin BL enterotoxin, a ternary complex of three proteins, is the causative agent of food poisoning and requires all three components for virulence. The X-ray structure of the binding domain of HBL suggests that it may form a pore similar to other soluble channel forming proteins. A putative pathway of pore formation is discussed.

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    • "Structurally, Hbl-B has a striking resemblance to HlyE/ClyA/SheA, a single-component hemolytic enterotoxin of the gram-negative bacteria Escherichia coli, Salmonella enterica, and Shigella flexneri [24]. This similarity is also predicted for the Nhe cellular binding proteins NheB and NheC [16], indicating that both gram-positive and gram-negative bacteria may have convergently evolved a similar pore-forming toxin. "
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    ABSTRACT: Bacillus cereus is a spore-forming, Gram-positive bacterium commonly associated with outbreaks of food poisoning. It is also known as an opportunistic pathogen causing clinical infections such as bacteremia, meningitis, pneumonia, and gas gangrene-like cutaneous infections, mostly in immunocompromised patients. B. cereus secretes a plethora of toxins of which four are associated with the symptoms of food poisoning. Two of these, the non-hemolytic enterotoxin Nhe and the hemolysin BL (Hbl) toxin, are predicted to be structurally similar and are unique in that they require the combined action of three toxin proteins to induce cell lysis. Despite their dominant role in disease, the molecular mechanism of their toxic function is still poorly understood. We report here that B. cereus strain ATCC 10876 harbors not only genes encoding Nhe, but also two copies of the hbl genes. We identified Hbl as the major secreted toxin responsible for inducing rapid cell lysis both in cultured cells and in an intraperitoneal mouse toxicity model. Antibody neutralization and deletion of Hbl-encoding genes resulted in significant reductions of cytotoxic activity. Microscopy studies with Chinese Hamster Ovary cells furthermore showed that pore formation by both Hbl and Nhe occurs through a stepwise, sequential binding of toxin components to the cell surface and to each other. This begins with binding of Hbl-B or NheC to the eukaryotic membrane, and is followed by the recruitment of Hbl-L1 or NheB, respectively, followed by the corresponding third protein. Lastly, toxin component complementation studies indicate that although Hbl and Nhe can be expressed simultaneously and are predicted to be structurally similar, they are incompatible and cannot complement each other.
    Full-text · Article · Oct 2013 · PLoS ONE
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    • "The three Nhe proteins share sequence homology with one another and also with the three Hbl proteins [12]. By using the crystal structure of Hbl-B [13] as a template, Fagerlund et al. (2008) constructed homology models of NheB and NheC which suggested that they could adopt similar structures. "
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    ABSTRACT: The structure of NheA, a component of the Bacillus cereus Nhe tripartite toxin, has been solved at 2.05 Å resolution using selenomethionine multiple-wavelength anomalous dispersion (MAD). The structure shows it to have a fold that is similar to the Bacillus cereus Hbl-B and E. coli ClyA toxins, and it is therefore a member of the ClyA superfamily of α-helical pore forming toxins (α-PFTs), although its head domain is significantly enlarged compared with those of ClyA or Hbl-B. The hydrophobic β-hairpin structure that is a characteristic of these toxins is replaced by an amphipathic β-hairpin connected to the main structure via a β-latch that is reminiscent of a similar structure in the β-PFT Staphylococcus aureus α-hemolysin. Taken together these results suggest that, although it is a member of an archetypal α-PFT family of toxins, NheA may be capable of forming a β rather than an α pore.
    Full-text · Article · Sep 2013 · PLoS ONE
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    • "Recently, homology modeling of NheB and NheC with one of the components of the related hemolysin of B. cereus, HblB, uncovered a striking similarity with the single-protein toxin cytolysin A (ClyA) (Fagerlund et al. 2008; Madegowda et al. 2008), despite weak sequence similarity. ClyA is an a-PFT produced by invasive enteric pathogens such as Escherichia coli Salmonella typhi and Shigella flexneri (Ludwig et al. 1995; Oscarsson et al. 2002). "
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    ABSTRACT: The nonhemolytic enterotoxin (Nhe) produced by Bacillus cereus is a pore-forming toxin consisting of three components, NheA, -B and -C. We have studied effects of Nhe on primate epithelial cells (Vero) and rodent pituitary cells (GH(4)) by measuring release of lactate dehydrogenase (LDH), K(+) efflux and the cytosolic Ca(2+) concentration ([Ca(2+)](i)). Plasma membrane channel events were monitored by patch-clamp recordings. Using strains of B. cereus lacking either NheA or -C, we examined the functional role of the various components. In both cell types, NheA + B + C induced release of LDH and K(+) as well as Ca(2+) influx. A specific monoclonal antibody against NheB abolished LDH release and elevation of [Ca(2+)](i). Exposure to NheA + B caused a similar K(+) efflux and elevation of [Ca(2+)](i) as NheA + B + C in GH(4) cells, whereas in Vero cells the rate of K(+) efflux was reduced by 50% and [Ca(2+)](i) was unaffected. NheB + C had no effect on either cell type. Exposure to NheA + B + C induced large-conductance steps in both cell types, and similar channel insertions were observed in GH(4) cells exposed to NheA + B. In Vero cells, NheA + B induced channels of much smaller conductance. NheB + C failed to insert membrane channels. The conductance of the large channels in GH(4) cells was about 10 nS. This is the largest channel conductance reported in cell membranes under quasi-physiological conditions. In conclusion, NheA and NheB are necessary and sufficient for formation of large-conductance channels in GH(4) cells, whereas in Vero cells such large-conductance channels are in addition dependent on NheC.
    Full-text · Article · Sep 2010 · Journal of Membrane Biology
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