Vasdev Kavita, Dhawan Shikha, Kapoor Rajeev Kumar, Kuhad Ramesh Chander
Department of Microbiology, University of Delhi South Campus, Benito Juarez Road, New Delhi 110021, India.
Fungal Genet Biol. 2005 Aug;42(8):684-93. doi: 10.1016/j.fgb.2005.03.013.
Cyathus bulleri, a bird's nest fungus, known to decolorize polymeric dye Poly R-478, was found to produce 8 U ml(-1) of laccase in malt extract broth. Laccase activity appeared as a single band on non-denaturing gel. Laccase was purified to homogeneity by anion exchange chromatography and gel filtration. The enzyme was a monomer with an apparent molecular mass of 60 kD, pI of 3.7 and was stable in the pH range of 2-6 with an optimum pH of 5.2. The optimal reaction temperature was 45 degrees C and the enzyme lost its activity above 70 degrees C. Enzyme could oxidize a broad range of various phenolic substrates. K(m) values for ABTS, 2,6-dimethoxyphenol, guaiacol, and ferulic acid were found to be 48.6, 56, 22, and 14 mM while K(cat) values were 204, 180, 95.6, and 5.2, respectively. It was completely inhibited by KCN, NaN(3), beta-mercaptoethanol, HgCl(2), and SDS, while EDTA had no effect on enzyme activity. The N-terminal amino acid sequence of C. bulleri laccase showed close homology to N-terminal sequences of laccase from other white-rot fungi. A 150 bp gene sequence encoding copper-binding domains I and II was most similar to the sequence encoding a laccase from Pycnoporus cinnabarinus with 74.8% level of similarity.
布氏鸟巢菌（Cyathus bulleri）是一种鸟巢菌，已知其能使聚合染料聚R - 478脱色，研究发现它在麦芽提取物肉汤中能产生8 U/ml的漆酶。漆酶活性在非变性凝胶上呈现为单一一条带。通过阴离子交换色谱和凝胶过滤将漆酶纯化至同质。该酶为单体，表观分子量为60 kD，pI为3.7，在pH 2 - 6范围内稳定，最适pH为5.2。最佳反应温度为45℃，酶在70℃以上失去活性。该酶能氧化多种不同的酚类底物。发现其对ABTS、2,6 - 二甲氧基苯酚、愈创木酚和阿魏酸的K(m)值分别为48.6、56、22和14 mM，而K(cat)值分别为204、180、95.6和5.2。它完全被KCN、NaN₃、β - 巯基乙醇、HgCl₂和SDS抑制，而EDTA对酶活性无影响。布氏鸟巢菌漆酶的N端氨基酸序列与其他白腐真菌漆酶的N端序列显示出密切的同源性。编码铜结合结构域I和II的150 bp基因序列与编码朱红密孔菌漆酶的序列最为相似，相似度为74.8%。
