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  Preparation of Structurally Diverse Chiral Alcohols by Engineering Ketoreductase CgKR

Zheng, G.-W., Liu, Y.-Y., Chen, Q., Huang, L., Yu, H.-L., Lou, W.-Y., et al. (2017). Preparation of Structurally Diverse Chiral Alcohols by Engineering Ketoreductase CgKR. ACS Catalysis, 7(10), 7174-7181. doi:10.1021/acscatal.7b01933.

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 Creators:
Zheng, Gao-Wei1, Author
Liu, Yuan-Yang1, Author
Chen, Qi1, Author
Huang, Lei1, Author
Yu, Hui-Lei1, Author
Lou, Wen-Yong2, Author
Li, Chun-Xiu1, Author
Bai, Yun-Peng1, Author
Li, Aitao3, Author              
Xu, Jian-He1, Author
Affiliations:
1State Key Laboratory of Bioreactor Engineering, Shanghai Collaborative Innovation Center for Biomanufacturing, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China, ou_persistent22              
2Lab of Applied Biocatalysis, School of Food Science and Engineering, South China University of Technology, Guangzhou 510640, Guangdong, China, ou_persistent22              
3Research Department Reetz, Max-Planck-Institut für Kohlenforschung, Max Planck Society, ou_1445588              

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Free keywords: asymmetric reduction; biocatalysis; chiral alcohol; ketoreductase; protein engineering; substrate specificity
 Abstract: Ketoreductases are tools for the synthesis of chiral alcohols in industry. However, the low activity of natural enzymes often restricts their use in industrial applications. On the basis of computational analysis and previous reports, two residues (F92 and F94) probably affecting the activity of ketoreductase CgKR1 were identified. By tuning these two residues, the CgKR1-F92C/F94W variant was obtained that exhibited higher activity toward all 28 structurally diverse substrates examined than the wild-type enzyme. Among them, 13 substrates have a specific activity over 50 U mg–1 (54–775 U mg–1). Using CgKR1-F92C/F94W as a catalyst, five substrates at high loading (>100 g–1 L–1) were reduced completely in gram-scale preparative reactions. This approach provides accesses to pharmaceutically relevant chiral alcohols with high enantioselectivity (up to 99.0% ee) and high space-time yield (up to 583 g–1 L–1 day–1). Molecular dynamics simulations highlighted the crucial role of residues 92 and 94 in activity improvement. Our findings provide useful guidance for engineering other ketoreductases, especially those possessing a similar active pocket to that in CgKR1.

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Language(s): eng - English
 Dates: 2017-06-142017-09-132017-10-06
 Publication Status: Published online
 Pages: 8
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: DOI: 10.1021/acscatal.7b01933
 Degree: -

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Title: ACS Catalysis
  Abbreviation : ACS Catal.
Source Genre: Journal
 Creator(s):
Affiliations:
Publ. Info: Washington, D.C. : American Chemical Society
Pages: - Volume / Issue: 7 (10) Sequence Number: - Start / End Page: 7174 - 7181 Identifier: Other: 2155-5435
CoNE: https://pure.mpg.de/cone/journals/resource/2155-5435