Permeabilization of model lipid membranes by Bacillus sphaericus mosquitocidal binary toxin and its individual components

The high larvicidal effect of Bacillus sphaericus (Bs), a mosquito control agent, originates from the presence of a binary toxin (Bs Bin) composed of two proteins (BinA and BinB) that work together to lyse gut cells of suscep tible larvae. We demonstrate for the first time that the binary toxin and i ts individual components permeabilize receptor-free large unilamellar phosp holipid vesicles (LUVs) and planar lipid bilayers (PLBs) by a mechanism of pore formation. Calcein-release experiments showed that LUV permeabilizatio n was optimally achieved at alkaline pH and in the presence of acidic lipid s. BinA was more efficient than BinB, BinB facilitated the BinA effect, and their stoichiometric mixture was more effective than the full Bin toxin. I n PLBs, BinA formed voltage-dependent channels of approximate to 100-200 pS with long open times and a high open probability. Larger channels (greater than or equal to 400 pS) were also observed. BinB, which inserted less eas ily, formed smaller channels (less than or equal to 100 pS) with shorter me an open times. Channels observed after sequential addition of the two compo nents, or formed by their 1:1 mixture (w/w), displayed BinA-like activity. Bs Bin toxin was less efficient at forming channels than the BinA/BinB mixt ure, with channels displaying the BinA channel behavior. Our data support t he concept of BinA being principally responsible for pore formation in lipi d membranes with BinB, the binding component of the toxin, playing a role i n promoting channel activity.