title: Purification and partial characterization of a fibrinolytic enzyme from the fruiting body of the medicinal and edible mushroom Pleurotus ferulae.
abstract: A fibrinolytic metalloprotease with in vitro fibrinolytic effects was purified from the edible mushroom Pleurotus ferulae using several chromatography steps including anion and ion exchange, gel filtration, and fast protein liquid chromatography columns. The molecular mass of the enzyme was estimated to be 20.0 kDa, as determined using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fibrin zymography. The protease was active at 50°C, and pH 4.0, 5.0, and 8.0. The fibrinolytic activity of the enzyme was inhibited by ethyleneglycol-bis-(2-aminoethyl)-N,N,N',N' tetraacetic acid and strongly inhibited by two metal ions, Cu and Mg. In vitro assays evaluating fibrinolytic activity on a fibrin plate, fibrin turbidity, and thrombolytic activity on fibrin clots using human fibrinogen and human thrombin revealed that the enzyme could hydrolyze fibrin polymers directly and inhibit the formation of fibrin clots. In activated partial thromboplastin time (APTT) and prothrombin time assays, the enzyme strongly prolonged the APTT, which detects an activity of intrinsic and common pathways. The enzyme showed strong in vivo protective effect against mortality/paralysis from epinephrine plus collagen-induced acute thromboembolism in in vivo model. Our findings suggest that the enzyme may have a potential for treatment and prevention of thrombosis-relative diseases.
authors.affiliation: a Department of Biomedical Science and BK21-Plus Research Team for Bioactive Control Technology , Gwangju , South Korea.
authors.forename: Jun-Hui
authors.initials: JH
authors.lastname: Choi
authors.affiliation: a Department of Biomedical Science and BK21-Plus Research Team for Bioactive Control Technology , Gwangju , South Korea.
authors.forename: Dae-Won
authors.initials: DW
authors.lastname: Kim
authors.affiliation: b Department of Bio-Health Science , Gwangju University , Gwangju , South Korea.
authors.forename: Seung
authors.initials: S
authors.lastname: Kim
authors.affiliation: a Department of Biomedical Science and BK21-Plus Research Team for Bioactive Control Technology , Gwangju , South Korea.
authors.forename: Sung-Jun
authors.initials: SJ
authors.lastname: Kim
mesh_terms: D000818:Animals; D001777:Blood Coagulation / Q000187:drug effects; D005343:Fibrinolytic Agents / Q000302:isolation & purification* / Q000378:metabolism / Q000627:therapeutic use; D048690:Fruiting Bodies, Fungal / Q000201:enzymology* / Q000378:metabolism; D006801:Humans; D008297:Male; D045726:Metalloproteases / Q000302:isolation & purification* / Q000378:metabolism / Q000627:therapeutic use; D008813:Mice, Inbred ICR; D020076:Pleurotus / Q000201:enzymology* / Q000378:metabolism; D013923:Thromboembolism / Q000097:blood / Q000188:drug therapy
chemical_list: D005343:Fibrinolytic Agents; D045726:Metalloproteases
keywords: Activated partial thromboplastin time; Pleurotus ferulae; anticoagulation; antithrombotic; fibrinogenolytic activity; metalloprotease