Proteolytic cleavage of Pseudomonas aeruginosa exotoxin A in the presence of lipid bilayers of different composition.

ABSTRACT

Pseudomonas aeruginosa exotoxin A (ETA) must be proteolytically nicked by furin at Arg279 before being translocated into the cytosol of target cells. A similar cleavage can also be obtained with trypsin. Using this assay we could show that the interaction with lipid bilayers can strongly influence the extent of nicking. We found that in the presence of vesicles containing negatively charged lipids ETA is cleaved into its two fragments A and B at enzyme concentrations approximately 50 times lower, or at pH values higher by 1.5 units, than in the absence of lipids. We suggest that the interaction with the lipid bilayer of the positively charged loop containing Arg279 provides the energy for its partial unfolding and makes it more accessible for proteolysis.