Preparation of Combined Cross-linked Enzyme Aggregates and Its Application in Synthesis of Chiral Alcohols by the Asymmetric Reduction of Carbanyl Group†

doi:10.7503/cjcu20170509

Chem. J. Chinese Universities ›› 2018, Vol. 39 ›› Issue (1): 54.doi: 10.7503/cjcu20170509

• Organic Chemistry • Previous Articles Next Articles

Preparation of Combined Cross-linked Enzyme Aggregates and Its Application in Synthesis of Chiral Alcohols by the Asymmetric Reduction of Carbanyl Group^†

YANG Meng¹, JIANG Huijuan¹, NING Chenxi², WEI Dongzhi², SU Erzheng^1,^2,^*()

1. College of Light Industry and Food Engineering, Nanjing Forestry University, Nanjing 210037, China
2. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China

Received:2017-07-26 Online:2018-01-10 Published:2017-11-28
Contact: SU Erzheng E-mail:ezhsu@njfu.edu.cn
Supported by:
† Supported by the National Natural Science Foundation of China(No.31401679), the Six Talent Peaks Project in Jiangsu Province, China(No.2015-JY-016), the China Postdoctoral Science Foundation(Nos.2016M600417, 2017T100373), the 333 Project of Jiangsu Province, China(No.BRA2017458) and the Open Funding Project of the State Key Laboratory of Bioreactor Engineering, China

Abstract

Abstract:

Using ketoreductase KRED 30 from Corynebacterium sp. and the D-glucose dehydrogenase from Bacillus subtilis as the model enzymes, a combined cross-linked enzyme aggregates(combi-CLEAs) with high activity was prepared by optimization of the precipitant type, glutaraldehyde concentration, cross-linking temperature and time. Characterization of the combi-CLEAs showed that the stability and substrate tolerance of free enzymes mixture were significantly improved after combined cross-linking. Investigation of catalytical performance showed that combi-CLEAs could effectively catalyze the reduction of ethyl 4-chloroacetoacetate(COBE) and 3'-chloroacetophenone(CPO) for the synthesis of corresponding chiral alcohols in aqueous system. Only a little cofactor was needed to start the reactions. The catalytical activity could retain more than 70% after continuously catalyzing for 10 batches. The result showed that catalytical performance of combi-CLEAs in the biphasic system was much better than that in aqueous system. The total turnover number(TTN) for catalyzing COBE could attend 6595, and TTN for catalyzing CPO could attend 7500. Thus, cofactor regeneration by combi-CLEAs is feasible.

Key words: Ketoreductase, D-Glucose dehydrogenase, Cofactor regeneration, Combined cross-linked enzyme aggregates(Combi-CLEAs), Chiral alcohol

TrendMD:

YANG Meng, JIANG Huijuan, NING Chenxi, WEI Dongzhi, SU Erzheng. Preparation of Combined Cross-linked Enzyme Aggregates and Its Application in Synthesis of Chiral Alcohols by the Asymmetric Reduction of Carbanyl Group^†[J]. Chem. J. Chinese Universities, 2018, 39(1): 54.

Figures/Tables 7

Scheme 1 Schematic view for the preparation of combined cross-linked enzyme aggregates and application in cofactor regeneration

Fig.1 Effect of glutaraldehyde content on the catalytical activity of the prepared combi-CLEAs during the cross-linking process

Fig.2 Effects of cross-linking time and temperature on the catalytical activity of the prepared combi-CLEAs during the cross-linking process

Fig.3 SEM images of combi-CLEAs in a less-structured form with different scale bar

Fig.4 Effects of substrate concentration on the catalytic activity of combi-CLEAs(a) and free enzymes mixture(b) in aqueous system(A, C) and biphasic system(B, D)

Fig.5 Reusability of combi-CLEAs catalyzed the reduction of ethyl 4-chloro-3-oxobutanoate(A) and 3'-chloroacetophenone(B) in aqueous system

Fig.6 Time courses for free enzymes mixture(A, C), combi-CLEAs(B, D) catalyzed the reduction of ethyl 4-chloro-3-oxobutanoate(A, B), 3'-chloroacetophenone(C, D) in biphasic system

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Preparation of Combined Cross-linked Enzyme Aggregates and Its Application in Synthesis of Chiral Alcohols by the Asymmetric Reduction of Carbanyl Group^†

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