Characterization of extracellular proteases of Aeromonas hydrophila

Two endoproteases, proteinase-I and proteinase-II, and one aminopeptidase were isolated from the culture supernatant of Aeromonas hydrophila by ammonium sulfate precipitation, gradient elution on diethylaminoethyl-Sephadex A-50 and Sephadex G-75 column chromatography. The molecular weights of the proteinases and aminopeptidase ranged between 30,000 to 48,000. The proteinases had comparable pH (8.5) and temperature (48~50°C) optima. The aminopeptidase assayed using L-leucine-ρ-nitroanilide showed maximum activity at pH 8.0 and at a temperature of 70°C. The aminopeptidase was remarkably heat stable, while the proteinases were heat sensitive. Both the proteinases hydrolysed several proteins including fish myofibrillar proteins. Divalent cations like Fe 2+ , Cu 2+ and Zn 2+ inhibited the activities of the two proteinases while Ca 2+ and Mg 2+ did not affect either the activities of the proteinases or that of the aminopeptidase. The aminopeptidase was inhibited by metal chelating agents as well as thiol reagents.