Edible Mushroom Lentinula edodes as Source of a Novel Fibrinolytic Enzyme: Biochemical Characterization
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Lentinula edodes were fermented in liquids to create a new fibrinolytic enzyme. DEAE sepharose chromatography, gel filtering using Sephadex-G200, and fractional precipitation by ammonium sulphate were used to purify the crude fibrinolytic enzyme. Using SDS-PAGE, the isolated enzyme's apparent molecular mass was determined to be 44 kDa. The obtained enzyme's optimal temperature and pH were 42℃ and 7.5, respectively. The fibrinolytic enzyme was completely inhibited by Hg2+ and partially inhibited by Cu2+, Ni2+ and Al3+. In decreasing order, Zn2+, Mg2+, Ca2+, K+, and Fe2+ all significantly increased its performance. The enzyme action was decreased by EGTA and EDTA, proving to be a metalloprotease. The serine inhibitors aprotinin and PMSF also inhibited the enzyme. In vitro, the purified enzyme demonstrated high selectivity for fibrin as a substrate. A chymotrypsin-like protease is suggested via amidolytic action directed at artificial substrates, which demonstrated great specificity to the synthetic peptide N-Succinyl-Ala-Ala-Pro-PhepNA. Moreover, the enzyme was found to improve thrombin time (TT) prothrombin time (PT), and activated partial thromboplastin time (APTT), while decreasing fibrinogen (FIB) and prothrombin activity (PA) levels. According to these relevant work, the enzyme has tremendous opportunity for usage as a natural treatment for thrombotic diseases and for preventing them.
Abstract
Lentinula edodes were fermented in liquids to create a new fibrinolytic enzyme. DEAE sepharose chromatography, gel filtering using Sephadex-G200, and fractional precipitation by ammonium sulphate were used to purify the crude fibrinolytic enzyme. Using SDS-PAGE, the isolated enzyme's apparent molecular mass was determined to be 44 kDa. The obtained enzyme's optimal temperature and pH were 42℃ and 7.5, respectively. The fibrinolytic enzyme was completely inhibited by Hg2+ and partially inhibited by Cu2+, Ni2+ and Al3+. In decreasing order, Zn2+, Mg2+, Ca2+, K+, and Fe2+ all significantly increased its performance. The enzyme action was decreased by EGTA and EDTA, proving to be a metalloprotease. The serine inhibitors aprotinin and PMSF also inhibited the enzyme. In vitro, the purified enzyme demonstrated high selectivity for fibrin as a substrate. A chymotrypsin-like protease is suggested via amidolytic action directed at artificial substrates, which demonstrated great specificity to the synthetic peptide N-Succinyl-Ala-Ala-Pro-PhepNA. Moreover, the enzyme was found to improve thrombin time (TT) prothrombin time (PT), and activated partial thromboplastin time (APTT), while decreasing fibrinogen (FIB) and prothrombin activity (PA) levels. According to these relevant work, the enzyme has tremendous opportunity for usage as a natural treatment for thrombotic diseases and for preventing them