Ene-reductases allow regio- and stereoselective reduction of activated C=C double bonds at the expense of nicotinamide adenine dinucleotide cofactors [NAD(P)H]. Biological NAD(P)H can be replaced by synthetic mimics to facilitate enzyme screening and process optimization. The ene-reductase FOYE-1, originating from an acidophilic iron oxidizer, has been described as a promising candidate and is now being explored for applied biocatalysis. Biological and synthetic nicotinamide cofactors were evaluated to fuel FOYE-1 to produce valuable compounds. A maximum activity of (319.7±3.2) U mg-1 with NADPH or of (206.7±3.4) U mg-1 with 1-benzyl-1,4-dihydronicotinamide (BNAH) for the reduction of N-methylmaleimide was observed at 30 °C. Notably, BNAH was found to be a promising reductant but exhibits poor solubility in water. Different organic solvents were therefore assayed: FOYE-1 showed excellent performance in most systems with up to 20 vol% solvent and at temperatures up to 40 °C. Purification and application strategies were evaluated on a small scale to optimize the process. Finally, a 200 mL biotransformation of 750 mg (R)-carvone afforded 495 mg of (2R,5R)-dihydrocarvone (>95 % ee), demonstrating the simplicity of handling and application of FOYE-1.

中文翻译：

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通过热稳定和耐有机溶剂的烯还原酶不对称还原（R）-香芹酮。

烯还原酶允许在区域和立体上选择性还原活化的C = C双键，但要以烟酰胺腺嘌呤二核苷酸辅因子[NAD（P）H]为代价。可以用合成模拟物代替生物NAD（P）H，以促进酶筛选和工艺优化。源自嗜酸性铁氧化剂的烯还原酶FOYE-1被描述为有前途的候选物，目前正在探索用于应用生物催化。对生物和合成烟酰胺辅助因子进行了评估，以为FOYE-1提供燃料，以生产有价值的化合物。观察到在30℃时，NADPH的（319.7±3.2）U mg-1或1-苄基-1,4-二氢烟酰胺（BNAH）的（206.7±3.4）U mg-1的最大还原活性为30。 ℃。值得注意的是，发现BNAH是有前途的还原剂，但在水中的溶解性较差。因此，分析了不同的有机溶剂：FOYE-1在大多数系统中使用最多20％的溶剂和在最高40°C的温度下均表现出出色的性能。对纯化和应用策略进行了小规模评估，以优化流程。最终，通过750 mg（R）-香芹酮的200 mL生物转化，得到495 mg（2R，5R）-二氢香芹酮（> 95％ee），证明了FOYE-1的操作和应用简单。