10-Hydroxy-2-decenoic acid (10-HDA) is an α,β-unsaturated medium-chain carboxylic acid containing a terminal hydroxyl group. It has various unique properties and great economic value. We improved the two-step biosynthesis method of 10-HDA. The conversion rate of the intermediate product trans-2-decenoic acid in the first step of 10-HDA synthesis could reach 93.1 ± 1.3% by combining transporter overexpression and permeation technology strategies. Moreover, the extracellular trans-2-decenoic acid content was five times greater than the intracellular content when 2.0% (v/v) triton X-100 and 1.2% (v/v) tween-80 were each used. In the second step of 10-HDA synthesis, we regenerated NAD(P)H by overexpressing a glucose dehydrogenase with the P450 enzyme (CYP153A33/M228L-CPR_BM3) in Escherichia coli, improving the catalytic performance of the trans-2-decenoic acid terminal hydroxylation. Finally, the yield of 10-HDA was 486.5 mg/L using decanoic acid as the substrate with two-step continuous biosynthesis. Our research provides a simplified production strategy to promote the two-step continuous whole-cell catalytic biosynthesis of 10-HDA and other α,β-unsaturated carboxylic acid derivatives.

中文翻译：

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使用 NAD(P)H 再生 P450 系统和全细胞催化生物合成高效生物合成 10-羟基-2-癸烯酸

10-羟基-2-癸烯酸（10-HDA）是一种含有末端羟基的α,β-不饱和中链羧酸。它具有多种独特的性能和巨大的经济价值。我们改进了10-HDA的两步生物合成方法。通过结合转运蛋白过表达和渗透技术策略，10-HDA合成第一步中间产物反式-2-癸烯酸的转化率可以达到93.1±1.3%。此外，当分别使用2.0%（v/v）triton X-100和1.2%（v/v）tween-80时，细胞外反式-2-癸烯酸含量比细胞内含量高5倍。在10-HDA合成的第二步中，我们通过在大肠杆菌中过表达葡萄糖脱氢酶P450酶(CYP153A33/M228L-CPR _BM3 )来再生NAD(P)H，提高反式-2-癸烯酸的催化性能末端羟基化。最终，以癸酸为底物，通过两步连续生物合成，10-HDA的产率为486.5 mg/L。我们的研究提供了一种简化的生产策略，以促进 10-HDA 和其他 α,β-不饱和羧酸衍生物的两步连续全细胞催化生物合成。