一株产漆酶细菌的分离鉴定及酶学性质研究

Isolation, identification of a laccase-producing bacterial strain and enzymatic properties of the laccase

摘要:

【目的】分离获得产漆酶的细菌菌株, 研究漆酶的酶学性质并应用于染料脱色。【方法】利用含铜的富集培养基筛选产漆酶细菌; 通过形态特征、生理生化试验及16S rDNA序列分析等方法进行鉴定; 以丁香醛连氮为底物测定漆酶的酶学性质; 通过测定染料在最大吸收波长下吸光值的变化评价漆酶对染料的脱色效果。【结果】从森林土壤中筛选到一株漆酶高产菌株LS05, 初步鉴定为解淀粉芽孢杆菌(Bacillus amyloliquefaciens); 菌株LS05的芽孢漆酶以丁香醛连氮为底物的最适pH为6.6, 最适温度为70 °C; 该酶具有较好的稳定性, 经70 °C处理10 h或在pH 9.0条件下放置10 d后可保留活性。对抑制剂SDS和EDTA具有一定的抗性, 在碱性条件下可有效脱色不同的工业染料, RB亮蓝、活性黑和靛红1 h内的脱色率达93%以上。【结论】Bacillus amyloliquefaciens LS05的芽孢漆酶在高温和碱性条件下稳定性强, 相对于真菌漆酶具有更好的工业应用特性, 可有效用于工业染料废水的处理。

Abstract:

[Objective] The aim of this study was to screen laccase-producing bacterial strains and to investigate the enzymatic properties as well as decolorization ability of the laccase. [Methods] Enrichment medium supplemented with copper ions was used to isolate bacterial strains exhibiting laccase activity. The isolated strain was identified by morphology observation, physiological and biochemical tests and 16S rDNA sequence analysis. The enzymatic properties of laccase were investigated with syringaldazine as substrate. Dye decolorization ability of the laccase was tested by determining the change at maximum wavelength of synthetic dyes. [Results] A bacterial strain LS05 with high laccase activity was isolated from forest soil, and was identified as Bacillus amyloliquefaciens. The spore laccase of strain LS05 demonstrated optimum pH and temperature at pH 6.6 and 70 °C, respectively. It also showed high stability, retaining its activity after incubation at 70 °C for 10 h or at pH 9.0 for 10 d. Resistance towards SDS and EDTA was found for the spore laccase. The enzyme could efficiently decolorize different synthetic dyes at alkaline conditions. More than 93% of remazol brilliant blue R, reactive black 5 and indigo carmine were decolorized within 1 h. [Conclusion] The spore laccase of Bacillus amyloliquefaciens LS05 was highly stable at high temperature and alkaline pH, which was more advantageous in industrial application than fungal laccase. It showed high potential in treatment of industrial dye effluents.