title: Characterization of an alkaline proteinase of fish muscle.
abstract: 1. The alkaline proteinase showing pH optimum 8.0 from white croaker (Sciaena schlegeli) skeletal muscle was purified electrophoretically homogeneously (2000-fold) using a combination of DEAE-cellulose chromatography, hydroxylapatite chromatography and Ultrogel AcA 34 gel filtration. 2. It was stable for 1 hr at 50 degrees C. The molecular weight of the enzyme was estimated to be 430,000 by gel filtration, with the enzyme composed of four kinds of subunits, the chain molecular weights of which were 45,000, 48,000, 51,000 and 57,000. 3. From the effects of inhibitors, the enzyme was identified as cysteine proteinase. ATP and Cu2+ inhibited the activity 50% at 10 mM and 70% at 0.1 mM, respectively. 4. Thus the enzyme was characterized as a high molecular weight, heat-stable, alkaline cysteine proteinase (HAP). 5. The enzyme showed hardly any activity below 50 degrees C but considerable activity at around 60 degrees C against myofibrils, digesting myosin heavy chain, actin and tropomyosin. With the addition of 5 M urea the enzyme hydrolyzed myofibrils well at around 30 degrees C.
authors.affiliation: Department of Fisheries, Faculty of Agriculture, Kinki University, Higashi-Osaka, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Makinodan
authors.forename: Y
authors.initials: Y
authors.lastname: Yokoyama
authors.forename: M
authors.initials: M
authors.lastname: Kinoshita
authors.forename: H
authors.initials: H
authors.lastname: Toyohara
mesh_terms: D000818:Animals; D003546:Cysteine Endopeptidases / Q000302:isolation & purification* / Q000378:metabolism; D004795:Enzyme Stability; D005399:Fishes; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology*; D010447:Peptide Hydrolases / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D046911:Macromolecular Substances; D010447:Peptide Hydrolases; D003546:Cysteine Endopeptidases