Heterologous production of a temperature and pH-stable laccase from Bacillus vallismortis fmb-103 in Escherichia coli and its application |
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| Affiliation: | 1. College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan 430074, China;2. Key Laboratory of Oil Crops Biology of Ministry of Agriculture in China, Oil Crops Research Institute of Chinese Academy of Agricultural Sciences, Wuhan 430064, China;1. Extremophiles Laboratory, Department of Microbiology, School of Biology and Center of Excellence in Phylogeny of Living Organisms, College of Science, University of Tehran, Tehran, Iran;2. Department of Biochemistry, Faculty of Biological Science, Tarbiat Modares University, Tehran, Iran;1. Faculty of Chemistry, University of Belgrade, Studentski trg 12-16, Belgrade, Serbia;2. Institute of Chemistry, Technology and Metallurgy-Center of Chemistry, University of Belgrade, Studentski trg 12-16, Belgrade, Serbia;1. Bioengineering Laboratory, RIKEN Cluster for Pioneering Research (CPR), RIKEN, 2-1 Hirosawa, Wako-shi, Saitama, 351-0198, Japan;2. Bioplastic Research Team, RIKEN Center for Sustainable Resource Science (CSRS), RIKEN, 2-1 Hirosawa, Wako-shi, Saitama, 351-0198, Japan;3. Department of Life Science and Technology, Tokyo Institute of Technology, 2-12-1 Ookayama, Meguro-ku, Tokyo, 152-8550, Japan |
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| Abstract: | To enhance laccase yield, the laccase gene from Bacillus vallismortis fmb-103 was cloned and heterologously expressed in Escherichia coli BL21 (DE3) cells. The auto-induction strategy was applied during fermentation, and the process was controlled, as follows: Cu2+ was added when the optical density at 600 nm (OD600) was 0.3, the fermentation temperature was adjusted to 16 °C when the OD600 was 0.9, and fermentation was stopped after 50 h. The yield of recombinant laccase was up to 3420 U/L, as assayed by 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid). Recombinant laccase was purified 4.47-fold by heating for 10 min at 70 °C and dialyzing against 50–60% ammonium sulfate, retained more than 50% activity after 10 h at 70 °C, and demonstrated broad pH stability. Malachite green was efficiently degraded by recombinant laccase, especially in combination with mediators. These results provided a basis for the future application of recombinant laccase to malachite green degradation. |
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| Keywords: | Laccase Heterologous production Characterization Malachite green degradation |
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