• Title, Summary, Keyword: Phenoloxidase

Search Result 32, Processing Time 0.11 seconds

Characterization of Laccase Purified from Korean Trametes hirsuta S1 (한국산 흰구름버섯(Trametes hirsuta S1)으로부터 정제된 Laccase의 특성)

  • Lim, Hyung-Seon;Yoon, Kyung-Ha
    • The Korean Journal of Mycology
    • /
    • v.32 no.2
    • /
    • pp.112-118
    • /
    • 2004
  • Laccase produced by Trametes hirsuta S1 isolated from Korea was partially purified and characterized using ultrafiltration, anion exchange chromatography and affinity chromatography. The laccase was produced as the predominant extracellular enzyme during primary metabolism. Neither lignin peroxidase nor veratryl alcohol oxidase (VAO) were detected in the culture fluid. Addition of 2,5-xylidine enhanced 4-fold laccase production. Purified laccase was a single polypeptide having a molecular mass of approximately 66 kDa, as determined by SDS-polyacrylamide gel electrophoresis, and carbohydrate content of 12%. $K_{m}\;and\;V_{max}$ values for laccase with ABTS [2,2-azinobis (3-ethylbenzthiazoline 6-sulfonic acid)] as a substrate (Lineweaver-Burk plot) was determined to $51.2\;{\mu}M\;and\;56.8\;{\mu}mole$, respectively. The optimal pH for laccase activity was found to be 3.0. The enzyme was very stable for 1 hour at $50^{\circ}C$. Half-life ($t_{1/2}$) of the enzyme was about 20 min at $70^{\circ}C$. Spectroscopic analysis of purified enzyme indicated that the enzyme was typical of copper-containing protein. Substrate specificity and inhibitor studies for laccase also indicated to be a typical fungal laccase. The N-terminal amino acid sequence of the T. hirsuta S1 laccase showed 100% of homology to those of laccase from C. hirsutus.

Characterization of Laccase Purified from Korean Pycnoporus cinnabarinus SCH-3 (한국산 주걱송편버섯(Pycnoporus cinnabarinus) SCH-3로부터 정제 된 Laccase의 특성)

  • Park, Eun-Hye;Yoon, Kyung-Ha
    • The Korean Journal of Mycology
    • /
    • v.31 no.2
    • /
    • pp.59-66
    • /
    • 2003
  • Laccase produced by Pycnoporus cinnabarinus SCH-3 isolated from Korea was partially purified using ultrafiltration, anion exchange chromatography and affinity chromatography, The laccase was produced as the predominant extracellular phenoloxidase during primary metabolism. Neither lignin peroxidase nor manganese-dependent peroxidase were detected in the culture fluid. In order to examine the effect of inducers in laccase production, 2,5-xylidine was added in the culture of Pycnoporus cinnabarinus SCH-3. Addition of 2,5-xylidine enhanced 25-fold laccase production. Purified laccase was a single polypeptide having a molecular mass of approximately 66 kDa, as determined by SDS-polyacrylamide gel electrophoresis, and carbohydrate content of 9%. $K_{m}\;and\;V_{max}$ values for laccase with ABTS [2,2-azinobis (3-ethylbenzthiazoline 6-sulfonic acid)] as a substrate (Lineweaver-Burk plot) was determined to be $44.4{\mu}M\;and\;56.0{\mu}mole$, respectively. The optimal pH for laccase activity was found to be 3.0. The enzyme was very stable for 1 hour at $60{\circ}C$. Half-life ($t_{1/2}$) of the enzyme was about 10 min at $80{\circ}C$. Spectroscopic analysis of purified enzyme indicated that the enzyme was typical of copper-containing protein. Substrate specificity and inhibitor studies for laccase also indicated to be a typical fungal laccase. The N-terminal amino acid sequence of the P. cinnabarinus SCH-3 laccase showed 94% of homology to the N-terminal sequences of laccases from P. cinnabarinus PB and P. coccineus.