Different modes of membrane permeabilization by two RTX toxins: HlyA from Escherichia coli and CyaA from Bordetella pertussis
Abstract
This study clarifies the membrane disruption mechanisms of two bacterial RTX toxins: alpha-hemolysin (HlyA) from Escherichia coli and a homologous adenylate cyclase toxin (CyaA) from Bordetella pertussis. We employed a fluorescence requenching method using liposomes with encapsulated fluorescent dye/quencher pair ANTS/DPX.
We showed that both toxins induced a graded leakage of liposome content with different selectivities for DPX and ANTS. In contrast to HlyA, CyaA exhibited a higher selectivity for cationic quencher DPX, which increased with vesicle diameter.
While disrupting liposomes, CyaA caused a highly cation-selective leakage whereas its mutated form with decreased channel K+/Cl− selectivity due to two substitutions in a predicted transmembrane segment (CyaA-E509K + E516K) exhibited much lower selectivity. We concluded that the fluorescence requenching method in combination with different size of liposomes is a valuable tool for characterization of pore-forming toxins and their variants.