Tags

Type your tag names separated by a space and hit enter

Carbonyl reductase identification and development of whole-cell biotransformation for highly efficient synthesis of (R)-[3,5-bis(trifluoromethyl)phenyl] ethanol.
Microb Cell Fact 2016; 15(1):191MC

Abstract

BACKGROUND

(R)-[3,5-bis(trifluoromethyl)phenyl] ethanol [(R)-3,5-BTPE] is a valuable chiral intermediate for Aprepitant (Emend) and Fosaprepitant (Ivemend). Biocatalyzed asymmetric reduction is a preferred approach to synthesize highly optically active (R)-3,5-BTPE. However, the product concentration and productivity of reported (R)-3,5-BTPE synthetic processes remain unsatisfied.

RESULTS

A NADPH-dependent carbonyl reductase from Lactobacillus kefir (LkCR) was discovered by genome mining for reduction of 3,5-bis(trifluoromethyl) acetophenone (3,5-BTAP) into (R)-3,5-BTPE with excellent enantioselectivity. In order to synthesize (R)-3,5-BTPE efficiently, LkCR was coexpressed with glucose dehydrogenase from Bacillus subtilis (BsGDH) for NADPH regeneration in Escherichia coli BL21 (DE3) cells, and the optimal recombinant strain produced 250.3 g/L (R)-3,5-BTPE with 99.9% ee but an unsatisfied productivity of 5.21 g/(L h). Then, four different linker peptides were used for the fusion expression of LkCR and BsGDH in E. coli to regulate catalytic efficiency of the enzymes and improved NADPH-recycling efficiency. Using the best strain (E. coli/pET-BsGDH-ER/K(10 nm)-LkCR), up to 297.3 g/L (R)-3,5-BTPE with enantiopurity >99.9% ee was produced via reduction of as much as 1.2 M of substrate with a 96.7% yield and productivity of 29.7 g/(L h).

CONCLUSIONS

Recombinant E. coli/pET-BsGDH-ER/K(10 nm)-LkCR was developed for the bioreduction of 3,5-BTAP to (R)-3,5-BTPE, offered the best results in terms of high product concentration and productivity, demonstrating its great potential in industrial manufacturing of (R)-3,5-BTPE.

Authors+Show Affiliations

State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, 200237, China.State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, 200237, China.State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, 200237, China.State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, 200237, China.State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, 200237, China. jplin@ecust.edu.cn.State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, 200237, China.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

27835967

Citation

Chen, Kangling, et al. "Carbonyl Reductase Identification and Development of Whole-cell Biotransformation for Highly Efficient Synthesis of (R)-[3,5-bis(trifluoromethyl)phenyl] Ethanol." Microbial Cell Factories, vol. 15, no. 1, 2016, p. 191.
Chen K, Li K, Deng J, et al. Carbonyl reductase identification and development of whole-cell biotransformation for highly efficient synthesis of (R)-[3,5-bis(trifluoromethyl)phenyl] ethanol. Microb Cell Fact. 2016;15(1):191.
Chen, K., Li, K., Deng, J., Zhang, B., Lin, J., & Wei, D. (2016). Carbonyl reductase identification and development of whole-cell biotransformation for highly efficient synthesis of (R)-[3,5-bis(trifluoromethyl)phenyl] ethanol. Microbial Cell Factories, 15(1), p. 191.
Chen K, et al. Carbonyl Reductase Identification and Development of Whole-cell Biotransformation for Highly Efficient Synthesis of (R)-[3,5-bis(trifluoromethyl)phenyl] Ethanol. Microb Cell Fact. 2016 Nov 11;15(1):191. PubMed PMID: 27835967.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Carbonyl reductase identification and development of whole-cell biotransformation for highly efficient synthesis of (R)-[3,5-bis(trifluoromethyl)phenyl] ethanol. AU - Chen,Kangling, AU - Li,Kefei, AU - Deng,Jian, AU - Zhang,Baoqi, AU - Lin,Jinping, AU - Wei,Dongzhi, Y1 - 2016/11/11/ PY - 2016/05/12/received PY - 2016/10/28/accepted PY - 2016/11/13/entrez PY - 2016/11/12/pubmed PY - 2017/4/5/medline KW - (R)-[3,5-bis(trifluoromethyl)phenyl] ethanol KW - 3,5-bis(trifluoromethyl) acetophenone KW - Carbonyl reductase KW - Fusion-protein expression KW - Glucose dehydrogenase SP - 191 EP - 191 JF - Microbial cell factories JO - Microb. Cell Fact. VL - 15 IS - 1 N2 - BACKGROUND: (R)-[3,5-bis(trifluoromethyl)phenyl] ethanol [(R)-3,5-BTPE] is a valuable chiral intermediate for Aprepitant (Emend) and Fosaprepitant (Ivemend). Biocatalyzed asymmetric reduction is a preferred approach to synthesize highly optically active (R)-3,5-BTPE. However, the product concentration and productivity of reported (R)-3,5-BTPE synthetic processes remain unsatisfied. RESULTS: A NADPH-dependent carbonyl reductase from Lactobacillus kefir (LkCR) was discovered by genome mining for reduction of 3,5-bis(trifluoromethyl) acetophenone (3,5-BTAP) into (R)-3,5-BTPE with excellent enantioselectivity. In order to synthesize (R)-3,5-BTPE efficiently, LkCR was coexpressed with glucose dehydrogenase from Bacillus subtilis (BsGDH) for NADPH regeneration in Escherichia coli BL21 (DE3) cells, and the optimal recombinant strain produced 250.3 g/L (R)-3,5-BTPE with 99.9% ee but an unsatisfied productivity of 5.21 g/(L h). Then, four different linker peptides were used for the fusion expression of LkCR and BsGDH in E. coli to regulate catalytic efficiency of the enzymes and improved NADPH-recycling efficiency. Using the best strain (E. coli/pET-BsGDH-ER/K(10 nm)-LkCR), up to 297.3 g/L (R)-3,5-BTPE with enantiopurity >99.9% ee was produced via reduction of as much as 1.2 M of substrate with a 96.7% yield and productivity of 29.7 g/(L h). CONCLUSIONS: Recombinant E. coli/pET-BsGDH-ER/K(10 nm)-LkCR was developed for the bioreduction of 3,5-BTAP to (R)-3,5-BTPE, offered the best results in terms of high product concentration and productivity, demonstrating its great potential in industrial manufacturing of (R)-3,5-BTPE. SN - 1475-2859 UR - https://www.unboundmedicine.com/medline/citation/27835967/Carbonyl_reductase_identification_and_development_of_whole_cell_biotransformation_for_highly_efficient_synthesis_of__R__[35_bis_trifluoromethyl_phenyl]_ethanol_ L2 - https://microbialcellfactories.biomedcentral.com/articles/10.1186/s12934-016-0585-5 DB - PRIME DP - Unbound Medicine ER -