Contribution of Cathepsin L to Autolysis of Sea Cucumber Stichopus japonicus Intestines

Abstract

ABSTRACTIn order to provide the fundamental basis for control and application of sea cucumber autolysis, the contribution of cathepsin L to intestinal proteolysis in sea cucumber (Stichopus japonicus) was investigated. Cathepsin L mRNA expression was significantly up-regulated by 1.80 ± 0.44-fold with ultraviolet irradiation (p< 0.05). According to the protein pattern identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and determination of the release of trichloroacetic acid (TCA)-soluble oligopeptide, high intestinal protein degradation was observed at pH 4.4 and 48°C. Intestinal protein proteolysis was strongly inhibited by cathepsin L inhibitors, including trans-epoxysuccinyl-L-leucyl-amido (4-guani-dino)butane (E-64), iodoacetic acid (IA), and antipain (AP). These results imply that cathepsin L is involved in intestinal autolysis in the sea cucumber S. japonicus.Abbreviations: TCA: trichloroacetic acid; Z-Phe-Arg-AMC: Z-Phe-Arg-7-amido-4-methylcou-marin; IA: iodoacetic acid; E-64: trans-epoxysuccinyl-L-leucyl-amido (4-guanidino) butane; AP: antipain; SBTI: soybean trypsin inhibitor; SDS-PAGE: sodium dodecyl sulfate polyacrylamide gel electrophoresis; RT-PCR: reverse transcription-polymerase chain reaction