title: Purification and characterization of collagenolytic proteases from the hepatopancreas of northern shrimp (Pandalus eous).
abstract: Three gelatinolytic proteases (A1, A2, and B) were purified using a synthetic substrate, DNP-Pro-Gln-Gly-Ile-Ala-Gly-Gln-d-Arg, from the hepatopancreas of Northern shrimp (Pandalus eous) by several chromatographic steps involving hydroxyapatite column chromatography, gel filtration on Superdex75, and ion-exchange chromatography on a MonoQ column. Collagenolytic proteases A2 and B, but not protease A1, were demonstrated to digest native porcine type I collagen at 25 degrees C and pH 7.5. Further characterizations of these two collagenolytic proteases showed that the pH optimum of enzyme A2 against DNP-peptide was found to be 11, whereas that of enzyme B was 8.5. The optimum temperature ranged between 40 and 45 degrees C for both enzymes, although enzyme B appeared to be thermally more stable than enzyme A2 at pH 7.5. Both enzymes were strongly inhibited by PMSF and antipain, which suggests that they belong to collagenolytic serine proteases.
authors.affiliation: Research & Development Center, Nichirei Corporation, 9 Shinminato, Mihama, Chiba, Chiba 261-8545, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Aoki
authors.forename: Md Nazmul
authors.initials: MN
authors.lastname: Ahsan
authors.forename: Kenji
authors.initials: K
authors.lastname: Matsuo
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Hagiwara
authors.forename: Shugo
authors.initials: S
authors.lastname: Watabe
mesh_terms: D000818:Animals; D002845:Chromatography; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D017364:Collagenases / Q000302:isolation & purification* / Q000378:metabolism*; D004064:Digestive System / Q000201:enzymology*; D017886:Durapatite; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D061965:Matrix Metalloproteinase Inhibitors; D033542:Pandalidae; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D013379:Substrate Specificity
chemical_list: D004791:Enzyme Inhibitors; D061965:Matrix Metalloproteinase Inhibitors; D015842:Serine Proteinase Inhibitors; D017886:Durapatite; D017364:Collagenases