Overexpression and secretion of proaerolysin by Aeromonas salmonicida by Sarah Ellen Burr.
Date
2001
Authors
Burr, Sarah Ellen
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Abstract
Aerolysin, a pore-forming toxin produced by Aeromonas species, is secreted as an inactive protoxin (proaerolysin) via the General Secretion Pathway. Recombinant PCR was used to generate proaerolysin variants containing internal deletions in key regions of the protein. This provided a means by which to study the role these protein domains play in proaerolysin secretion. Three proaerolysin variants could not be secreted by A. salmonicida and were shown to fold incorrectly within the cell.
Overexpression of proaerolysin by A. salmonicida resulted in an increase in toxin secretion into the extracellular medium and an accumulation of toxin within the cells. Further examination carried out using pulse-chase analysis indicated that the cells contain two distinct pools of proaerolysin; one that is rapidly secreted by the cell and a second that is not readily available for secretion. Electron microscopic examination of A. salmonicida indicated that intracellular proaerolysin accumulates within an enlarged periplasm. The results obtained suggest that there is a limit to the amount of protein that can be efficiently exported from the cell via the General Secretion Pathway.