期刊
BIORESOURCE TECHNOLOGY
卷 101, 期 17, 页码 6761-6767出版社
ELSEVIER SCI LTD
DOI: 10.1016/j.biortech.2010.03.099
关键词
Construction; Co-expression; Shine-Dalgarno regions; Aligned spacing; Biocatalysis
资金
- Major Basic Research Program of China [2009CB724700]
- National Key Technology RD Program [2008BAI63B07]
- Innovation Fund for Doctoral Dissertation of Nanjing University of Technology
Biocatalysis of ethyl 4-chloro-3-oxobutanoate (CUBE) to ethyl (S)-4-chloro-3-hydroxybutanoate [(S)-CHBE] was carried out using Escherichia coli co-expressing a carbonyl reductase gene from Pichia stipitis and a glucose dehydrogenase gene from Bacillus megaterium. An efficient polycistronic plasmid with a high-level of enzyme co-expression was constructed by changing the order of the genes, altering the Shine-Dalgarno (SD) regions, and aligned spacing (AS) between the SD sequence and the translation initiation codon. The optimal SD sequence was 5-TAAGGAGG-3, and the optimal AS distance was eight nucleotides. Asymmetric reduction of COBE to (S)-CHBE with more than 99% enantiomeric excess was demonstrated by transformants, using a water/ethyl caprylate system. The recombinant cells produced 1260 mM product in the organic phase, and the total turnover number, defined as moles (S)-CHBE formed per mole NADP(+), was 12,600, which was more than 10-fold higher than in aqueous systems. (C) 2010 Elsevier Ltd. All rights reserved.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据