While lignocellulose is a promising source of renewable sugars for microbial fermentations, the presence of inhibitory compounds in typical lignocellulosic feedstocks, such as furfural, has hindered their utilisation. In Escherichia coli, a major route of furfural toxicity is the depletion of NADPH pools due to its use as a substrate by the YqhD enzyme that reduces furfural to its less toxic alcohol form. Here, we examine the potential of exploiting benzyl alcohol dehydrogenases as an alternative means to provide this same catalytic function but using the more abundant reductant NADH, as a strategy to increase the capacity for furfural removal. We determine the biochemical properties of three of these enzymes, from Pseudomonas putida, Acinetobacter calcoaceticus, and Burkholderia ambifaria, which all demonstrate furfural reductase activity. Furthermore, we show that the P. putida and B. ambifaria enzymes are able to provide substantial increases in furfural tolerance in vivo, by allowing more rapid conversion to furfuryl alcohol and resumption of growth. The study demonstrates that methods to seek alternative cofactor dependent enzymes can improve the intrinsic robustness of microbial chassis to feedstock inhibitors.

中文翻译：

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由异源 NADH 依赖性苯甲醇脱氢酶介导的大肠杆菌中糠醛耐受性的改善

虽然木质纤维素是用于微生物发酵的可再生糖的有前途的来源，但在典型的木质纤维素原料（例如糠醛）中存在抑制性化合物阻碍了它们的利用。在大肠杆菌中，糠醛毒性的主要途径是NADPH池的消耗，因为它被YqhD酶用作底物，可将糠醛还原为其毒性较小的醇形式。在这里，我们研究了利用苯甲醇脱氢酶作为提供相同催化功能的替代方法的潜力，但使用更丰富的还原剂 NADH 作为提高糠醛去除能力的策略。我们确定了其中三种酶的生化特性，它们来自恶臭假单胞菌、醋酸钙不动杆菌和伯克霍尔德菌，它们都表现出糠醛还原酶的活性。此外，我们表明恶臭假单胞菌和 B. ambifaria 酶能够通过允许更快地转化为糠醇和恢复生长来显着增加体内糠醛耐受性。该研究表明，寻找替代辅因子依赖性酶的方法可以提高微生物底盘对原料抑制剂的内在稳定性。