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A loop engineering strategy improves laccase lcc2 activity in ionic liquid and aqueous solution


Laccases and especially high redox potential laccase play an important role in lignin degradation. The fungal laccase lcc2 from Trametes versicolor has a high redox potential and EMIM- and BMIM-based ionic liquids show excellent solubilization of wooden biomass. Concentrations of EMIM- and BMIM to efficiently dissolve lignin impede laccase activity. Protein engineering to improve the activity and resistance of laccases in ionic liquids offers a promising opportunity for lignin valorization for the sustainable production of fuels and bulk high-value chemicals. In this work, we have performed computational assisted protein engineering of lcc2 to increase performance in presence of ionic liquid and aqueous solution. We showed that the loop L1 (amino acids residues 284-320) is highly important for improving lcc2 activity in EMIM EtSO4 and aqueous solutions. Lcc2 activity was improved based on a KnowVolution campaign through site saturation of seven amino acid positions identified by computational modeling. Simultaneous site saturation of four amino acid positions by OmniChange yielded variants OM1 (A285P/A310R/A312E/A318G) and OM3 (A310D/A312P/A318R) with a 3.9-fold (535.8± 36.9 U/mg) and 1.6-fold (216.8± 5.3 U/mg) increased specific activity in aqueous solution (lcc2 WT, 138.9± 6.5 U/mg), respectively. High conservation of the loop L1 in fungal laccases suggests that computational assisted loop engineering might be used as a general strategy to improve the activity in ionic liquids and aqueous solutions.

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Publication details

The article was received on 17 Dec 2017, accepted on 04 May 2018 and first published on 04 May 2018

Article type: Paper
DOI: 10.1039/C7GC03776G
Citation: Green Chem., 2018, Accepted Manuscript
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    A loop engineering strategy improves laccase lcc2 activity in ionic liquid and aqueous solution

    A. M. Wallraf, H. Liu, L. Zhu, G. Khalfallaha, C. Simons, H. Alibiglou, M. D. Davari and U. Schwaneberg, Green Chem., 2018, Accepted Manuscript , DOI: 10.1039/C7GC03776G

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