A PORE-FORMING PROTEIN WITH A PROTEASE-ACTIVATED TRIGGER

Alpha-Hemolysin (alphaHL) is a 293 amino acid pore-forming toxin, which is secreted as a water-soluble monomer by Staphylococcus aureus. By forming a hexameric pore, alphaHL damages the plasma membranes of target cells. Previous studies established that alphaHL proteins with nicks near the midpoint of a central glycine-rich loop are held together by a domain - domain interaction and are hemolytically active. In contrast, alphaHL proteins comprising two alphaHL truncation mutants that overlap in the central loop have no or greatly reduced pore-forming activity, even though the two chains again form a tight complex. Based on these findings, overlap mutants have now been designed that are activated when redundant amino acids in the loop are removed by proteases. Further, the identity of the activating enzyme can be specified by additional mutagenesis of the protease recognition site in the overlap sequence. Mutants of alphaHL that are activated by tumor-associated proteases might be useful components of immunotoxins.