A novel multicopper oxidase (laccase) from cyanobacteria: Purification, characterization with potential in the decolorization of anthraquinonic dye

A novel extracellular laccase enzyme produced from Spirulina platensis CFTRI was purified by ultrafiltration, cold acetone precipitation, anion exchange and size exclusion chromatography with 51.5% recovery and 5.8 purification fold. The purified laccase was a monomeric protein with molecular mass of ~66 kDa that was confirmed by zymogram analysis and peptide mass fingerprinting. The optimum pH and temperature of the enzyme activity was found at 3.0 and 30°C using ABTS as substrate but the enzyme was quite stable at high temperature and alkaline pH. The laccase activity was enhanced by Cu+2, Zn+2 and Mn+2. In addition, the dye decolorization potential of purified laccase was much higher in terms of extent as well as time. The purified laccase decolorized (96%) of anthraquinonic dye Reactive blue- 4 within 4 h and its biodegradation studies was monitored by UV visible spectra, FTIR and HPLC which concluded that cyanobacterial laccase can be efficiently used to decolorize synthetic dye and help in waste water treatment.

从钝顶螺旋藻CFTRI株（Spirulina platensis CFTRI）中分离得到的新型胞外漆酶（laccase），经超滤、冷丙酮沉淀、阴离子交换色谱及尺寸排阻色谱纯化后，回收率达51.5%，纯化倍数为5.8倍。纯化所得漆酶为单体蛋白，分子量约66 kDa，该结论经酶谱分析（zymogram analysis）与肽质量指纹图谱（peptide mass fingerprinting）验证。以ABTS（2,2'-联氮-双(3-乙基苯并噻唑啉-6-磺酸)）为底物时，该酶的最适反应pH为3.0，最适反应温度为30℃；但该酶在高温及碱性pH环境下仍具备良好稳定性。Cu²+、Zn²+与Mn²+可显著提升该漆酶的酶活性。此外，纯化漆酶的染料脱色性能在脱色程度与反应时长两方面均表现突出：其可在4小时内将蒽醌类染料活性蓝-4（Reactive blue 4）脱色96%。通过紫外-可见光谱、傅里叶变换红外光谱（FTIR）及高效液相色谱（HPLC）对该漆酶的生物降解过程进行监测分析后证实，蓝藻来源漆酶可高效实现合成染料脱色，可为废水处理工作提供有力支撑。