Abstract: In this paper, the enzymatic properties of two aspartic proteases, cathepsin D (Cat D) and cathepsin E (Cat E), in sea cucumber were studied, and their involvement in the autolysis of sea cucumber was also investigated. Crude enzyme was extracted from the body wall of sea cucumbers with Tris-HCl buffer (pH 7.0). The specific substrate fluorescence method was used to determine the enzymatic properties of Cat D and Cat E. Results showed that Cat D and Cat E exhibited maximum activities at pH 5.0 and 60 ℃, and pH 4.0 and 40 ℃, respectively. Their activities kept stable at 20–40 ℃. Zn2+, Cu2+, Fe3+, and Mn2+ inhibited Cat D activity by 86%, 76.3%, 29.2%, 56.5% and 48.5%, respectively. Fe3+, Fe2+ and Cu2+ inhibited Cat E by 99.1%, 82.2% and 28.6%, respectively. Pepstatin A, Z-LLL, PMSF, and 1,10-phenanthroline inhibited the two proteases by 98%–99%, 65%–78%, 30%–35%, and 19%–23%, respectively. L-Cys, DDT and EDTA increased their activities by 30.4%–31.1%, 7.58%–9.64%, and 6.6%–7.9%, respectively. These results suggested that Cat D and Cat E were sensitive to metal ions and they were metal ion-dependent aspartic proteases, with cysteine and serine around the active center involved in modulating the enzymatic activities. Moreover, they might be involved in protein degradation during the autolysis of sea cucumber.

Key words: Stichopus japonicus, aspartic proteases, cathepsin D, cathepsin E, enzymatic property, autolysis