Secretion of Hemolysin of Aeromonas sobria as Protoxin and Contribution of the Propeptide Region Removed from the Protoxin to the Proteolytic Stability of the Toxin

Abstract

The sequence at the amino terminus region of the hemolysin of Aeromonas sobria is homologous with that of aerolysin of A. hydrophila. However, there is no homology between the two toxins in the sequence at the carboxy terminal region. It has been shown that aerolysin is secreted into culture supernatant as aprotoxin. This proaerolysin is activated by the proteolytic removal of a carboxy terminal peptide. However, the role of the carboxy terminal region, which is removed in the activation process, has not been elucidated.In this study, we showed that hemolysin is also secreted as a protoxin into culture supernatant and that prohemolysin is cleaved by the protease of A. sobria between Ser-446 and Ala-447, resulting in the removal of a 42 amino acid peptide. The removal of the peptide converts the prohemolysin into active hemolysin. Subsequently, we mutated the hemolysin gene to delete the last several amino acid residues and expressed the genes in Escherichia coli, in order to examine the role of the carboxy terminal region of prohemolysin. The amounts of these mutant hemolysins accumulated in the periplasmic space of E. coli were very low compared with that of the wild-type. This observation indicated that the carboxy terminal region of prohemolysin contributes to the proteolytic stability of the toxin.

Salt in Surroundings Influences the Production of Serine Protease into Milieu by Aeromonas sobria

“…It has been reported that ALH emerges as a precursor in culture medium and then the precursor is processed to the active form by protease(s) released by the cell itself (8,9,21). In in vitro experiments, ALH can be processed by incubation with trypsin.…”

Aerolysin is activated by metalloprotease in Aeromonas veronii biovar sobria

“…Studies on the cleavage site of A. sobria aerolysin have been carried out by Nomura et al (1999). By determining the Cterminal region of mature aerolysin from the culture supernatant, the cleavage site of the toxin was determined to be between Ser446 and Ala447.…”

“…The toxin is secreted to the culture supernatant as a precursor (named proaerolysin), which has no biological activity. A C-terminal peptide must be removed to convert proaerolysin into active aerolysin (Nomura et al, 1999). Evidence has shown that proaerolysin can be activated by protease(s) produced by host cells (Abrami et al, 1998) or by Aeromonas itself (Howard & Buckley, 1985).…”

Fluid Secretion Caused by Aerolysin-Like Hemolysin of Aeromonas sobria in the Intestines Is Due to Stimulation of Production of Prostaglandin E 2 via Cyclooxygenase 2 by Intestinal Cells

“…We purified and characterized the enterotoxin from the culture supernatant of A. sobria isolated from a patient with diarrhea and demonstrated that it possessed hemolytic activity in addition to enterotoxic activity (14,26).…”