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Enhancing the organic solvent resistance of transaminase from Aspergillus terreus by regional random mutation
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  • Chun-Ning Wang,
  • Shuai Qiu,
  • Fang-Fang Fan,
  • Chang-Jiang Lyu,
  • Sheng Hu,
  • Wei-Rui Zhao,
  • Jia-Qi Mei,
  • Le-He Mei,
  • Jun Huang
Chun-Ning Wang
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Fang-Fang Fan
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Chang-Jiang Lyu
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Wei-Rui Zhao
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Jia-Qi Mei
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Jun Huang

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Biocatalysis in high-concentration organic solvents has been applied to produce various industrial products with many advantages. However, using enzymes in organic solvents often suffers from inactivation or decreased catalytic activity and stability. So, improving the tolerance of enzymes in organic solvents is essential. Herein, the method of regional random mutation combined with combinatorial mutation was used to improve the resistance of transaminase from Aspergillus terreus (AtATA) in organic solvents, and the best mutant T23I/T200K/P260S (M3) was acquired. In different concentrations of dimethyl sulfoxide (DMSO), the catalytic efficiency toward 1-acetylnaphthalene and the stability were higher than the wild-type (WT) of AtATA. M3 also showed enhanced stability against six organic solvents with different oil-water partition coefficients (log P values). The results of decreased Root Mean Square Fluctuation (RMSF) values via 20-ns molecular dynamics simulations under different concentration DMSO revealed that mutant M3 had lower flexibility, acquiring a more stable protein structure and contributing to its organic solvents stability than WT. Intra- and intermolecular interaction analysis indicated that the increased hydrogen bonds and hydrophobic interactions within monomers or at the interface of two monomers also strengthened the stability of the overall structure against organic solvents. Furthermore, M3 was applied to convert 1-acetylnaphthalene for synthesizing (R)-(+)-1(1-naphthyl)-ethylamine ((R)-NEA), which was and an intermediate of Cinacalcet Hydrochloride. Moreover, 3~10 mM 1-acetylnaphthalene can be converted to (R)-NEA with 94.2~38.9% yield and a strict R-stereoselectivity within 10 h under 25% DMSO, which was higher than WT and expected to be a potential biocatalyst for industrial application.
13 Mar 2023Submitted to Biotechnology Journal
14 Mar 2023Submission Checks Completed
14 Mar 2023Assigned to Editor
17 Mar 2023Reviewer(s) Assigned
14 Apr 2023Review(s) Completed, Editorial Evaluation Pending
14 Apr 2023Editorial Decision: Revise Major
09 May 20231st Revision Received
10 May 2023Assigned to Editor
10 May 2023Submission Checks Completed
10 May 2023Reviewer(s) Assigned
23 May 2023Review(s) Completed, Editorial Evaluation Pending
23 May 2023Editorial Decision: Revise Major
29 May 20232nd Revision Received
29 May 2023Submission Checks Completed
29 May 2023Assigned to Editor
01 Jun 2023Reviewer(s) Assigned
12 Jun 2023Review(s) Completed, Editorial Evaluation Pending
14 Jun 2023Editorial Decision: Accept