A molecular pin to study the dynamics of β-barrel formation in pore-forming toxins on erythrocytes: a sliding model

Abstract.

γ-Hemolysins are pore-forming toxins which develop from water-soluble monomers by combining two different ‘albeit homologous’ proteins. They form oligomeric pores in both cell and model membranes by undergoing a still poorly understood conformational rearrangement in the stem region. The stem is formed by three β-strands, folded onto the core of the soluble protein and completely extended in the pore. We propose a new model to explain such a process. Seven double-cysteine mutants were developed by inserting one cysteine on the stretch that links the β-hairpin to the core of the protein and another on different positions along the β-strands. The membrane bound protein was blocked in a non-lytic state by S–S bond formation. Six mutants were oxidized as inactive intermediates, but became active after adding DTT. These results demonstrate that the stem extension can be temporarily frozen and that the β-barrel formation occurs by β-strand concerted step-by-step sliding.