Study on a prolyl endopeptidase from the skeletal muscle of common carp (Cyprinus carpio) |
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Authors: | Meng-Xiang Wang Chan Zhong Qiu-Feng Cai Guang-Ming Liu Ling Zhang Kenji Hara Wen-Jin Su Min-Jie Cao |
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Affiliation: | 1. College of Biological Engineering, The Key Laboratory of Science and Technology for Aquaculture and Food Safety, Jimei University, Xiamen 361021, China;2. Faculty of Fisheries, Nagasaki University, Nagasaki 852-8521, Japan |
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Abstract: | A prolyl endopeptidase (PEP) was purified to homogeneity from the skeletal muscle of common carp using a procedure involving ammonium sulfate fractionation and column chromatography involving DEAE-Sephacel, Phenyl-Sepharose, DEAE-Sepharose Fast Flow, and hydroxyapatite. The molecular weight of the PEP was 82 kDa as determined by SDS-PAGE. Using Suc-Gly-Pro-MCA as a substrate, the optimal pH and temperature of the purified enzyme were pH 6.0 and 35 °C, respectively, and the Km and kcat were 8.33 μM and 1.71 S?1, respectively. The activity of the PEP was inhibited by SUAM-14746, a specific inhibitor of prolyl endopeptidases, and was partially inhibited by the serine proteinase inhibitors PMSF and Pefabloc SC. According to peptide mass fingerprinting, 12 peptide fragments with a total of 134 amino acid residues were obtained, which were highly identical to prolyl endopeptidases from zebrafish (Danio rerio) and sponge (Amphimedon queenslandica), confirming the purified enzyme was a prolyl endopeptidase. Our present study for the first time reported the existence of a prolyl endopeptidase in fish muscle. |
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