Efficient production of (R)-2-hydroxy-4-phenylbutyric acid by using a coupled reconstructed D-lactate dehydrogenase and formate dehydrogenase system.
Efficient production of (R)-2-hydroxy-4-phenylbutyric acid by using a coupled reconstructed D-lactate dehydrogenase and formate dehydrogenase system.
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DOI：
10.1371/journal.pone.0104204
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Xu P
中科院分区：
文献类型：
--
作者：
Sheng B;Zheng Z;Lv M;Zhang H;Qin T;Gao C;Ma C;Xu P
(R)-2-羟基-4-苯基丁酸 [(R)-HPBA] 是生产血管紧张素转换酶抑制剂的关键前体。然而，已报道的(R)-HPBA合成方法的产物收率和浓度仍然不能令人满意。保加利亚乳杆菌 ATCC 11842 中 NAD 依赖性 d-乳酸脱氢酶 (d-nLDH) 的 Y52L/F299Y 突变体被发现对 2-氧代-4-苯基丁酸 (OPBA) 具有高生物还原活性。然后将突变体 d-nLDHY52L/F299Y 与大肠杆菌 BL21 (DE3) 中的甲酸脱氢酶共表达，构建新型生物催化剂大肠杆菌 DF。因此，开发了一种新的生物还原工艺，利用大肠杆菌 DF 的全细胞作为生物催化剂，甲酸盐作为辅因子再生的共底物，用于从 OPBA 生产 (R)-HPBA。然后优化生物催化条件。在最佳条件下，73.4 mM OPBA 在 90 分钟内还原为 71.8 mM (R)-HPBA。鉴于其高产物对映体过量（> 99％）和生产率（47.9 mM h−1），构建的偶联生物催化系统是（R）-HPBA生产的有前途的替代方案。
(R)-2-Hydroxy-4-phenylbutyric acid [(R)-HPBA] is a key precursor for the production of angiotensin-converting enzyme inhibitors. However, the product yield and concentration of reported (R)-HPBA synthetic processes remain unsatisfactory. The Y52L/F299Y mutant of NAD-dependent d-lactate dehydrogenase (d-nLDH) in Lactobacillus bulgaricus ATCC 11842 was found to have high bio-reduction activity toward 2-oxo-4-phenylbutyric acid (OPBA). The mutant d-nLDHY52L/F299Y was then coexpressed with formate dehydrogenase in Escherichia coli BL21 (DE3) to construct a novel biocatalyst E. coli DF. Thus, a novel bio-reduction process utilizing whole cells of E. coli DF as the biocatalyst and formate as the co-substrate for cofactor regeneration was developed for the production of (R)-HPBA from OPBA. The biocatalysis conditions were then optimized. Under the optimum conditions, 73.4 mM OPBA was reduced to 71.8 mM (R)-HPBA in 90 min. Given its high product enantiomeric excess (>99%) and productivity (47.9 mM h−1), the constructed coupling biocatalysis system is a promising alternative for (R)-HPBA production.
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