The properties of Bacillus cereus hemolysin II pores depend on environmental conditions |
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Authors: | Zhanna I Andreeva Maria G Fomkina Natalia E Suzina Alexander S Solonin |
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Institution: | a Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences, Pushchino, Moscow Region, 142290, Russia b Institute of Theoretical and Experimental Biophysics, Russian Academy of Sciences, Moscow Region, Pushchino, 142290, Russia c Institute of Cell Biophysics, Russian Academy of Sciences, Moscow Region, Pushchino, 142290, Russia d State Research Center of Virology and Biotechnology “VECTOR”, Koltsovo, Novosibirsk Region, 630559, Russia |
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Abstract: | Hemolysin II (HlyII), one of several cytolytic proteins encoded by the opportunistic human pathogen Bacillus cereus, is a member of the family of oligomeric β-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 °C and 4 °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. |
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Keywords: | αHL α-hemolysin of Staphylococcus aureus HlyII hemolysin II of Bacillus cereus RBC red blood cells HU hemolytic unit βPFT β-barrel pore-forming toxin PMSF phenylmethylsulfonylfluoride PEG polyethylene glycol |
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