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KMID : 0545120100200071069
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Journal of Microbiology and Biotechnology
2010 Volume.20 No. 7 p.1069 ~ p.1076
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Purification and characterization of a laccase from the edible wild mushroom Tricholoma mongolicum
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Miao Li
Guoqing Zhang
Hexiang Wang
Tzibun Ng
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Abstract
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A novel laccase from Tricholoma mongolicum was purified by using a procedure which entailed ion exchange chromatography on DEAE-cellulose, CM-cellulose, Q-Sepharose, and FPLC-gel filtration on Superdex 75. The purified enzyme was obtained with a specific activity of 1480 U (mg protein)-1 and a final yield of 15%. It was found to be a monomeric protein with a molecular mass of 66 kDa as estimated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Its N-terminal amino acid sequence was GIGPVADLYVGNRIL, similar to some but different other mushroom laccase. The optimum pH and temperature for the purified enzyme were pH 2 to pH 3 and 30 °C, respectively. It displayed a low Km toward 2,7′-azinobis (3-ethylbenzothiazolone-6-sulfonic acid) diammonium salt (ABTS) and high Kcat/Km values. The purified laccase oxidized a wide range of lignin-related phenols, but exerted maximal activity on ABTS. It was significantly inhibited by Hg2+ ions, and remarkably stimulated by Cu2+ ions. It inhibited HIV-1 reverse transcriptase and proliferation of hepatoma HepG2 cells and breast cancer MCF7 cells with an IC50 of 0.65 μM, 1.4 μM, and 4.2 μM, respectively, indicating that it is also an antipathogenic protein.
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KEYWORD
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Laccase, Tricholoma mongolicum, Purification, Characterization
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