Abstract: In this paper, serine protease in the body wall of sea cucumber (Stichopus japonicus) was investigated on its partial properties and role in the autolysis of sea cucumber. The serine protease was obtained after a series of processes including extraction with Tris-HCl buffer, (NH4)2SO4 precipitation, dialysis, and ultrafiltration. Results showed that the maximum protease activity was observed using Boc-Phe-Ser-Arg-MCA as substrate. Based on this substrate, the peak protease activity occurred at pH 6.2 and 8.5 and 50–55 ℃. The protease activity could be significantly inhibited by Cu2+, Fe3+, Pb2+ and Zn2+ but activated by Ca2+. The activity was partially inhibited by phenanthroline, EDTA, and cysteine-modifying reagents, while it together with the production of soluble proteins and trichloroacetic acid (TCA)-soluble peptides as a result of autolysis could be significantly inhibited by 4-(2-aminoethyl)benzenesulfonyl fluoride hydrochloride (AEBSF), phenylmethanesulfonyl fluoride (PMSF), N-p-tosyl-L-phenylalanine chloromethyl ketone (TPCK), (3S)-1-chloro-3-tosylamido-7-amino-2-heptanone hydrochloride (TLCK), and soybean trypsin inhibitor (SBTI). The above results suggest that there are at least two metal ion-dependent trypsin/trypsin-like serine proteases in the body wall of S. japonicus, with cysteine being involved in the active center or substrate-identifying site, and these proteases may be involved in the autolysis of S. japonicus.

Key words: sea cucumber, trypsin and trypsin-like serine protease, characterization, autolysis