Two-Enzyme Coexpressed Recombinant Strain for Asymmetric Synthesis of Ethyl (R)-2-Hydroxy-4-phenylbutyrate

doi:10.1016/S1872-2067(11)60436-1

Abstract

Abstract: (R)-2-Hydroxy-4-phenylbutyrate (HPBE) is an important chiral intermediate for the synthesis of angiotensin-converting enzyme (ACE) inhibitors. Asymmetric reduction of ethyl 2-oxo-4-phenyl-butyrate (OPBE) to (R)-HPBE using a r ecombinant strain can provide high enantioselectivity. Cofactor regeneration is a critical issue in the application of a recombinant strain. A carbonyl reductase gene (iolS) and a glucose dehydrogenase (GDH) gene from Bacillus subtilis were cloned. Recombinant IolS was purified using a Ni-NTA column and its enzyme activity properties were investigated. The purified IolS exhibited maximum activity at pH 6.0 and 30 ^oC, and the enzyme showed good thermostability below 40 ^oC. It retained over 75% of its activity in the acidic pH range of 5.5-7.0. Three coexpression strategies were used for the recombinant vectors. The recombinant E. coli strain containing polycistronic plasmid pET-G-T7-I showed excellent carbonyl reductase activity, and the specific activity of both IolS and GDH in the crude cell extract reached 1.5 U/mg. In the asymmetric reduction of OPBE by recombinant E. coli cells in aqueous system, the yield of (R)-HPBE reached over 99% with an enantiomeric excess of 99.5% at 10 g/L of OPBE within 15 h.

Key words: carbonyl reductase, glucose dehydrogenase, coexpression, (R)-2-hydroxy-4-phenylbutyrate, asymmetric reduction

SU Yu-Ning, NI Ye, WANG Jun-Chao, XU Zhi-Hao, SUN Zhi-Hao. Two-Enzyme Coexpressed Recombinant Strain for Asymmetric Synthesis of Ethyl (R)-2-Hydroxy-4-phenylbutyrate[J]. Chinese Journal of Catalysis, 2012, 33(10): 1650-1660.

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URL: https://www.cjcatal.com/EN/10.1016/S1872-2067(11)60436-1

https://www.cjcatal.com/EN/Y2012/V33/I10/1650

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