Purification and characteristics of trypsin from masu salmon (<Emphasis Type="Italic">Oncorhynchus masou</Emphasis>) cultured in fresh-water |
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| Authors: | Gaku Kanno Takahito Yamaguchi Hideki Kishimura Etsurou Yamaha Hiroki Saeki |
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| Institution: | (1) Laboratory of Marine Products and Food Science, Research Faculty of Fisheries Sciences, Hokkaido University, Hokkaido 041-8611, Japan;(2) Nanae Fresh-Water Laboratory, Field Science Center for Northern Biosphere, Hokkaido University, Hokkaido 041-8611, Japan; |
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| Abstract: | Trypsin from the pyloric ceca of masu salmon (Oncorhynchus masou) cultured in fresh water was purified by a series of chromatographies including Sephacryl S-200, Sephadex G-50 and diethylaminoethyl
cellulose to obtain a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS–PAGE) and native PAGE.
The molecular mass of the purified trypsin was estimated to be approximately 24,000 Da by SDS–PAGE. The enzyme activity was
strongly inhibited by phenylmethylsulfonyl fluoride, soybean trypsin inhibitor, and N
α
-p-tosyl-l-lysine chloromethyl ketone. Masu salmon trypsin was stabilized by calcium ion. The optimum pH of the masu salmon trypsin
was around pH 8.5, and the trypsin was unstable below pH 5.0. The optimum temperature of the masu salmon trypsin was around
60°C, and the trypsin was stable below 50°C, like temperate-zone and tropical-zone fish trypsins. The N-terminal 20 amino acid sequence of the masu salmon trypsin was IVGGYECKAYSQPHQVSLNS, and its charged amino acid content was
lower than those of trypsins from frigid-zone fish and similar to those of trypsins from temperate-zone and tropical-zone
fish. In the phylogenetic tree, the masu salmon trypsin was classified into the group of the temperate-zone fish trypsin. |
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