The first fungal laccase with an alkaline pH optimum obtained by directed evolution and its application in indigo dye decolorization
2019 | journal article. A publication with affiliation to the University of Göttingen.
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The first fungal laccase with an alkaline pH optimum obtained by directed evolution and its application in indigo dye decolorization
Yin, Q.; Zhou, G.; Peng, C.; Zhang, Y.; Kües, U. ; Liu, J. & Xiao, Y. et al. (2019)
AMB Express, 9(1). DOI: https://doi.org/10.1186/s13568-019-0878-2
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- Authors
- Yin, Qiang; Zhou, Gang; Peng, Can; Zhang, Yinliang; Kües, Ursula ; Liu, Juanjuan; Xiao, Yazhong; Fang, Zemin
- Abstract
- Abstract Engineering of fungal laccases with optimum catalytic activity at alkaline pH has been a long-lasting challenge. In this study, a mutant library containing 3000 clones was obtained by error-prone PCR to adapt the optimum pH of a fungal laccase Lcc9 from the basidiomycete Coprinopsis cinerea . After three rounds of functional screening, a mutant with three amino acid changes (E116K, N229D, I393T) named PIE5 was selected. PIE5 showed an optimum pH of 8.5 and 8.0 against guaiacol and 2,6-DMP when expressed in Pichia pastoris , representing the first fungal laccase that possesses an optimum pH at an alkaline condition. Site directed mutagenesis disclosed that N229D contributed the most to the optimum pH increment. A single N229D mutation caused an increase in optimum pH by 1.5 units. When used in indigo dye decolorization, PIE5 efficiently decolorized 87.1 ± 1.1% and 90.9 ± 0.3% indigo dye at the optimum conditions of pH 7.0–7.5 and 60 °C, and with either methyl 3,5-dimethoxy-4-hydroxybenzoate or 2,2′-azino-bis(3-ethylbenzothazoline-6-sulfonate) as the mediator. In comparison, the commercially available fungal laccase TvLac from Trametes villosa decolorized 84.3 ± 1.8% of indigo dye under its optimum conditions (opt. pH 5.0 and 60 °C). The properties of an alkaline-dependent activity and the high indigo dye decolorization ability (1.3-fold better than the parental Lcc9) make the new fungal laccase PIE5 an alternative for specific industrial applications.
- Issue Date
- 2019
- Journal
- AMB Express
- eISSN
- 2191-0855
- Language
- English