title: Purification and characteristics of trypsin from masu salmon (Oncorhynchus masou) cultured in fresh-water.
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 ( alpha )-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 degrees C, and the trypsin was stable below 50 degrees 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.
authors.affiliation: Laboratory of Marine Products and Food Science, Research Faculty of Fisheries Sciences, Hokkaido University, Hokkaido, 041-8611, Japan.
authors.forename: Gaku
authors.initials: G
authors.lastname: Kanno
authors.affiliation: Laboratory of Marine Products and Food Science, Research Faculty of Fisheries Sciences, Hokkaido University, Hokkaido, 041-8611, Japan.
authors.forename: Takahito
authors.initials: T
authors.lastname: Yamaguchi
authors.affiliation: Laboratory of Marine Products and Food Science, Research Faculty of Fisheries Sciences, Hokkaido University, Hokkaido, 041-8611, Japan. kishi@fish.hokudai.ac.jp.
authors.forename: Hideki
authors.initials: H
authors.lastname: Kishimura
authors.affiliation: Nanae Fresh-Water Laboratory, Field Science Center for Northern Biosphere, Hokkaido University, Hokkaido, 041-8611, Japan.
authors.forename: Etsurou
authors.initials: E
authors.lastname: Yamaha
authors.affiliation: Laboratory of Marine Products and Food Science, Research Faculty of Fisheries Sciences, Hokkaido University, Hokkaido, 041-8611, Japan.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Saeki
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002845:Chromatography; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007421:Intestine, Small / Q000201:enzymology*; D007564:Japan; D008969:Molecular Sequence Data; D017683:Oncorhynchus / Q000378:metabolism*; D010664:Phenylmethylsulfonyl Fluoride / Q000378:metabolism; D010802:Phylogeny; D020539:Sequence Analysis, Protein; D013696:Temperature; D014357:Trypsin / Q000737:chemistry / Q000302:isolation & purification*; D014361:Trypsin Inhibitors / Q000378:metabolism
chemical_list: D014361:Trypsin Inhibitors; D010664:Phenylmethylsulfonyl Fluoride; D014357:Trypsin
references.citation: FEBS Lett. 1975 Aug 1;56(1):139-43
references.pmid: 1157929
references.citation: Comp Biochem Physiol B Biochem Mol Biol. 2006 May;144(1):47-56
references.pmid: 16500127
references.citation: Nature. 1970 Aug 15;227(5259):680-5
references.pmid: 5432063
references.citation: Comp Biochem Physiol B. 1984;79(4):613-22
references.pmid: 6518765
references.citation: Food Chem. 2008 Sep 15;110(2):352-60
references.pmid: 26049226
references.citation: J Mol Biol. 1974 Feb 25;83(2):185-208
references.pmid: 4821870
references.citation: J Agric Food Chem. 2006 Jul 26;54(15):5617-22
references.pmid: 16848554
references.citation: J Agric Food Chem. 2007 May 30;55(11):4548-53
references.pmid: 17469841
references.citation: J Mol Evol. 1997 Dec;45(6):640-52
references.pmid: 9419241
references.citation: Comp Biochem Physiol B Biochem Mol Biol. 2007 Dec;148(4):382-9
references.pmid: 17707670
references.citation: Can J Biochem Physiol. 1959 Dec;37:1393-9
references.pmid: 14405350
references.citation: Nucleic Acids Res. 1994 Nov 11;22(22):4673-80
references.pmid: 7984417
references.citation: J Biol Chem. 1951 Nov;193(1):265-75
references.pmid: 14907713
references.citation: Comp Biochem Physiol B Biochem Mol Biol. 2005 Jan;140(1):91-8
references.pmid: 15621514
references.citation: Protein Eng. 1994 Jan;7(1):57-64
references.pmid: 8140095
references.citation: Gene. 1986;41(2-3):305-10
references.pmid: 3011602
references.citation: Biochemistry. 1973 Aug 14;12(17):3146-53
references.pmid: 4738933
references.citation: Comp Biochem Physiol B. 1988;91(4):677-84
references.pmid: 3224506
references.citation: Comp Biochem Physiol B Biochem Mol Biol. 2002 Jun;132(2):485-90
references.pmid: 12031475
references.citation: Eur J Biochem. 1995 Sep 1;232(2):677-85
references.pmid: 7556223
references.citation: Comp Biochem Physiol B. 1982;71(4):557-62
references.pmid: 7083813
references.citation: Biochemistry. 1977 Feb 22;16(4):654-64
references.pmid: 556951
references.citation: Comput Appl Biosci. 1996 Aug;12(4):357-8
references.pmid: 8902363
references.citation: Eur J Biochem. 1989 Mar 1;180(1):85-94
references.pmid: 2707266
references.citation: Biochim Biophys Acta. 1996 Nov 14;1298(1):45-57
references.pmid: 8948488