韦伯灵芝漆酶的分离纯化及其性质  

Purification and Characterization of Laccase from Ganoderma weberianum

作　　者：陈琼华[1] 周玉萍[2] 陈晓[1] 柯德森[1] 程惠贞[2] 田长恩[1,2] 

机构地区：[1]广州大学生命科学学院,广东广州510006 [2]广州大学植物抗逆基因功能研究广州市重点实验室,广东广州510006

出　　处：《食品科学》2010年第5期201-205,共5页Food Science

基　　金：广东省自然科学基金项目(8151009101000013);广东省科技计划项目(2007B080701008);广州市高校科学技术研究项目(62031);广州大学植物抗逆基因功能研究广州市重点实验室开放基金项目(200801)

摘　　要：采用硫酸铵分级盐析、透析、聚乙二醇浓缩和高效液相色谱柱层析从韦伯灵芝TZC-1发酵液中分离纯化得到电泳纯漆酶,其纯化倍数为37.1倍,酶活性回收率为21.3%。活性-PAGE(聚丙烯酰胺凝胶电泳)结果表明该漆酶由一种同工酶组成,纯化漆酶经SDS-PAGE检测显示为单一条带,其分子质量约为40kD。该漆酶催化底物ABTS的最适反应温度为50～60℃,最适反应pH值为4.6,在60℃以下和pH3.0～5.0范围内保持稳定,以ABTS为底物的表观Km值为13.8μmol/L。Fe2+完全抑制酶活,Al3+和Mn2+对该漆酶也有明显抑制作用,Hg+、Cu2+和Mg2+对该酶有明显激活作用,而Zn2+、Ba2+及K+对该酶活性影响不大。The laccase from Ganoderma weberianum TZC-1 was purified using fractional ammonium sulfate precipitation, dialysis, polyethylene glycol condensation and high performance liquid column chromatography. Compared to the crude extract, its purity exhibited a 37.1-fold improvement and the recovery rate of enzyme activity was 21.3%. Meanwhile, SDS-PAGE analysis revealed a single band with molecular weight of 40 kD. Moreover, the optimal reaction conditions of this laccase towards ABTS as substrate included 50-60 ℃ of reaction temperature and pH 4.6. The Km value was 13.8 μmol/L under these optimal reaction conditions. Furthermore, the activity of the enzyme exhibited excellent stability below 60 ℃ and pH 3.0 - 5.5, but could be inhibited by Fe2＋, Al3＋ and Mn2＋ and enhanced by Hg＋, Cu2＋ and Mg2＋. No obvious effect on laccase activity was observed due to Zn2＋, Ba2＋ and K＋.

关 键 词：韦伯灵芝 漆酶 纯化 酶活 

分 类 号：Q554[生物学—生物化学]