DIMERIZATION STABILIZES THE PORE-FORMING TOXIN AEROLYSIN IN SOLUTION

Aerolysin is a channel-forming protein secreted as a protoxin by Aerom onas hydrophila. Analytical centrifugation measurements showed that pr oaerolysin is a dimer in solution, and this was confirmed by chemical cross-linking with dimethyl suberimidate. Dissociation of proaerolysin with low concentrations of SDS resulted in the loss of tertiary struc ture, assessed by near ultraviolet circular dichroism. This was accomp anied by an increase in the protein's ability to bind the hydrophobic dye 1-anilino-8-naphthalene sulfonate, as well as by increased sensiti vity to proteolytic degradation. However, the monomer was not fully un folded by the detergent, as the tryptophans remained in a hydrophobic environment, and the secondary structure measured by far ultraviolet c ircular dichroism did not seem to be affected. Aerolysin, the active f orm of the protein, was also shown to be a dimer, and its stability wa s found to be no different from the stability of the protoxin dimer. S ubstituting tryptophan 371 or tryptophan 373 with leucine greatly redu ced the stability of dimeric proaerolysin. These substitutions are kno wn to increase the protein's ability to oligomerize, supporting the co nclusion that dimer dissociation is necessary for oligomerization to o ccur.