Insufficient rate of NADPH regeneration often limits the activity of biosynthetic pathways. Expression of NADPH-regenerating enzymes is commonly used to address this problem and increase cofactor availability. Here, we construct an Escherichia coli NADPH-auxotroph strain, which is deleted in all reactions that produce NADPH with the exception of 6-phosphogluconate dehydrogenase. This strain grows on a minimal medium only if gluconate is added as NADPH source. When gluconate is omitted, the strain serves as a “biosensor” for the capability of enzymes to regenerate NADPH in vivo. We show that the NADPH-auxotroph strain can be used to quantitatively assess different NADPH-regenerating enzymes and provide essential information on expression levels and concentrations of reduced substrates required to support optimal NADPH production rate. The NADPH-auxotroph strain thus serves as an effective metabolic platform for evaluating NADPH regeneration within the cellular context.

中文翻译：

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NADPH-营养缺陷型大肠杆菌：一种用于测试NADPH体内再生的传感器菌株

NADPH再生速率不足通常会限制生物合成途径的活性。NADPH再生酶的表达通常用于解决此问题并增加辅助因子的利用率。在这里，我们构建了一个大肠杆菌NADPH-营养缺陷型菌株，该菌株在除6-磷酸葡萄糖酸脱氢酶以外的所有产生NADPH的反应中均被删除。仅当添加葡萄糖酸盐作为NADPH来源时，该菌株才能在基本培养基上生长。如果省略了葡萄糖酸盐，则该菌株可作为“生物传感器”来检测酶在体内再生NADPH的能力。。我们表明，NADPH营养缺陷型菌株可用于定量评估不同的NADPH再生酶，并提供表达水平和支持最佳NADPH生成率所需的还原底物浓度的基本信息。因此，NADPH-营养缺陷型菌株可作为评估细胞内NADPH再生的有效代谢平台。