Guo Haipeng, Zheng Bingsong, Jiang Dean, Qin Wensheng
Department of Biology, Lakehead University, Thunder Bay, ON, Canada.
J Mol Microbiol Biotechnol. 2017;27(4):217-227. doi: 10.1159/000478859. Epub 2017 Sep 1.
Laccases from bacteria have been widely studied in the past 2 decades due to the higher growth rate of bacteria and their excellent thermal and alkaline pH stability. In this study, a novel laccase gene was cloned from Bacillus sp., analyzed, and functionally expressed in Escherichia coli. The laccase was highly induced in the E. coli expression system with a maximum intracellular activity of 16 U mg-1 protein. The optimal temperature and pH of the purified laccase were 40°C and 4.6, respectively, when ABTS (2,2'-azino-bis[3-ethylbenzothiazoline-6-sulfonate]) was used as the substrate. The purified laccase showed high stability in the pH range of 3.0-9.0, and retained more than 70% of its activity after 24 h of incubation at 40°C with a pH value of 9.0. Furthermore, the enzyme exhibited extremely high temperature and ion metal tolerance. The half-life of the purified laccase at 70°C was 15.9 h. The purified laccase could efficiently decolorize 3 chemical dyes, especially in the presence of ABTS as a mediator. The high production of this laccase in E. coli and exceptional characteristics of the recombinant enzyme protein make it a promising candidate for industrial applications.
在过去20年里，由于细菌生长速度较快且具有出色的热稳定性和碱性pH稳定性，细菌漆酶受到了广泛研究。在本研究中，从芽孢杆菌属中克隆了一个新型漆酶基因，进行了分析，并在大肠杆菌中实现了功能表达。该漆酶在大肠杆菌表达系统中被高度诱导，细胞内最大活性为16 U mg-1蛋白质。以ABTS（2,2'-联氮-双-（3-乙基苯并噻唑啉-6-磺酸））为底物时，纯化漆酶的最适温度和pH分别为40°C和4.6。纯化漆酶在pH值为3.0 - 9.0的范围内表现出高稳定性，在40°C、pH值为9.0的条件下孵育24小时后仍保留超过70%的活性。此外，该酶表现出极高的温度和离子金属耐受性。纯化漆酶在70°C下的半衰期为15.9小时。纯化漆酶能够高效地使3种化学染料脱色，尤其是在存在ABTS作为介体的情况下。这种漆酶在大肠杆菌中的高产量以及重组酶蛋白的优异特性使其成为工业应用的一个有前景的候选者。
