Ma Xiaojian, Liu Lu, Li Qingqing, Liu Yunyun, Yi Li, Ma Lixin, Zhai Chao
Hubei Collaborative Innovation Center for Green Transformation of Bio-Resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
Enzyme Microb Technol. 2017 Aug;103:34-41. doi: 10.1016/j.enzmictec.2017.04.004. Epub 2017 Apr 21.
Laccases are oxidoreductase catalyze the oxidation of a wide range of substrates with oxygen as the electron acceptor. This report was aimed to the high-level expression of a laccase, CueO from Escherichia coli K12 in Pichia pastoris GS115 and its application on decolorization of synthetic dyes. The yacK gene coding CueO was cloned into an expression vector of Pichia pastoris, pHBM905BDM and expressed in a secretory form with Pichia pastoris GS115 as the host. The yield of the recombinant protein was 556mg/L with high-density fermentation and the enzyme activity was about 41,000U/L. The recombinant laccase was purified and characterized. Its optimum pH and temperature was 3.0 and 55°C with 2, 2'-azino-bis-(3-ethylbenzothazoline-6-sulfonic acid) (ABTS) as the substrate, respectively. This recombinant protein was thermostable and its half life at 70°C was about 60min. In the presence of natural redox mediator acetosyringone, the purified recombinant laccase decolorized 98.1% and 98.5% of Congo red, malachite green, respectively. It also decolorized 90.03% of Remazol brilliant blue R without this mediator. In addition, this enzyme was applied on the decolorization of wastewater from a textile printing factory and showed an obvious bleaching effect.
漆酶是一种氧化还原酶，它以氧气作为电子受体催化多种底物的氧化反应。本报告旨在实现来自大肠杆菌K12的漆酶CueO在毕赤酵母GS115中的高水平表达及其在合成染料脱色中的应用。编码CueO的yacK基因被克隆到毕赤酵母表达载体pHBM905BDM中，并以毕赤酵母GS115作为宿主以分泌形式表达。通过高密度发酵，重组蛋白的产量为556mg/L，酶活性约为41,000U/L。对重组漆酶进行了纯化和表征。以2, 2'-联氮-双-(3-乙基苯并噻唑啉-6-磺酸)(ABTS)为底物时，其最适pH和温度分别为3.0和55°C。该重组蛋白具有热稳定性，在70°C下的半衰期约为60分钟。在天然氧化还原介质乙酰丁香酮存在下，纯化的重组漆酶分别使刚果红、孔雀石绿脱色98.1%和98.5%。在没有这种介质的情况下，它也能使活性艳蓝R脱色90.03%。此外，该酶应用于一家纺织印染厂废水的脱色，显示出明显的脱色效果。
