Bacillus cereus can attack the cell membranes of the alga Chara corallina by means of HlyII.

We studied the influence of Bacillus cereus bacteria on cells of the freshwater alga Chara corallina. These bacteria and recombinant Bacillus subtilis strains are capable of producing the secreted toxin HlyII, which changes the electrophysiological parameters of the algal electrically excitable plasma membrane by forming pores. Cooperative incubation of bacterial cells, which carry active hlyII gene, and Chara corallina cells caused a decrease in the resting potential (V(m)) and plasma membrane resistance (R(m)) of algal cells. The efficiency of each strain was commensurable with its ability to produce HlyII. Purified hemolysin II caused a similar effect on V(m) and R(m) of intact and perfused cells. This protein changed the kinetics and magnitude of transient voltage-dependent calcium and calcium-activated chloride currents owing to the formation of additional Ca(2+)-permeable pores in algal cell membrane. Occurrence of the cellulose cell wall with pores 2.1 to 4.6nm in diameter suggests that HlyII molecules reach the plasma membrane surface strictly as monomers.

Expression of Bacillus cereus hemolysin II in Bacillus subtilis renders the bacteria pathogenic for the crustacean Daphnia magna.

Hemolysin II (HlyII) is a pore-forming toxin of the opportunistic pathogen Bacillus cereus. Despite our understanding of the mechanism of HlyII cytotoxicity in vitro, many of its characteristics, including potential target cells, conditions of its action and expression, are not known. Here we report that the expression of hlyII in Bacillus subtilis renders the bacteria hemolytic and is able to kill the crustacean Daphnia magna. The hemolytic activity of hlyII-encoded B. subtilis strains in culture media is positively correlated with virulence in D. magna. Fluorescence microscopy reveals postinfection changes in the mitochondrial potential of intestinal tissue, suggesting that the formation of ionic pores leads to cell death. In the presence of the transcriptional regulator HlyIIR, HlyII expression decreases 200-fold, and B. subtilis expressing both hlyII and hlyIIR remains hemolytic, but not pathogenic to the crustacean.

The properties of Bacillus cereus hemolysin II pores depend on environmental conditions.

Hemolysin II (HlyII), one of several cytolytic proteins encoded by the opportunistic human pathogen Bacillus cereus, is a member of the family of oligomeric beta-barrel pore-forming toxins. This work has studied the pore-forming properties of HlyII using a number of biochemical and biophysical approaches. According to electron microscopy, HlyII protein interacts with liposomes to form ordered heptamer-like macromolecular assemblies with an inner pore diameter of 1.5-2 nm and an outer diameter of 6-8 nm. This is consistent with inner pore diameter obtained from osmotic protection assay. According to the 3D model obtained, seven HlyII monomers might form a pore, the outer size of which has been estimated to be slightly larger than by the other method, with an inner diameter changing from 1 to 4 nm along the channel length. The hemolysis rate has been found to be temperature-dependent, with an explicit lag at lower temperatures. Temperature jump experiments have indicated the pore structures formed at 37 degrees C and 4 degrees C to be different. The channels formed by HlyII are anion-selective in lipid bilayers and show a rising conductance as the salt concentration increases. The results presented show for the first time that at high salt concentration HlyII pores demonstrate voltage-induced gating observed at low negative potentials. Taken together we have found that the membrane-binding properties of hemolysin II as well as the properties of its pores strongly depend on environmental conditions. The study of the properties together with structural modeling allows a better understanding of channel functioning.

Sizing the pore of the volume-sensitive anion channel by differential polymer partitioning.

Partitioning of ethylene glycol and its polymeric forms into the pore of the volume-sensitive outwardly rectifying (VSOR) anion channel was studied to assess the pore size. Polyethylene glycol (PEG) PEG 200-300 (Rh = 0.27-0.53 nm) effectively suppressed the single-channel currents, whereas PEG 400-4000 (Rh = 0.62-1.91 nm) had little or no effect. Since all the molecules tested effectively decreased electric conductivity of the bulk solution, the observed differential effects between PEG 200-300 and PEG 400-4000 on the VSOR single-channel current are due to their limited partitioning into the channel lumen. The cut-off radius of the VSOR channel pore was assessed to be 0.63 nm.