Furan-2,5-dicarboxylic acid (FDCA) is a building block of biodegradable plastics that can be used to replace those derived from fossil carbon sources. In recent years, much interest has focused on the synthesis of FDCA from the bio-based 5-hydroxymethylfurfural (HMF) through a cascade of enzyme reactions. Aryl-alcohol oxidase (AAO) and 5-hydroxymethylfurfural oxidase (HMFO) are glucose-methanol-choline flavoenzymes that may be used to produce FDCA from HMF through three sequential oxidations, and without the assistance of auxiliary enzymes. Such a challenging process is dependent on the degree of hydration of the original aldehyde groups and of those formed, the rate-limiting step lying in the final oxidation of the intermediate 5-formyl-furancarboxylic acid (FFCA) to FDCA. While HMFO accepts FFCA as a final substrate in the HMF reaction pathway, AAO is virtually incapable of oxidizing it. Here, we have engineered AAO to perform the stepwise oxidation of HMF to FDCA through its structural alignment with HMFO and directed evolution. With a 3-fold enhanced catalytic efficiency for HMF and a 6-fold improvement in overall conversion, this evolved AAO is a promising point of departure for further engineering aimed at generating an efficient biocatalyst to synthesize FDCA from HMF.

中文翻译：

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通过释放的芳基醇氧化酶将5-羟甲基糠醛顺序氧化为呋喃2,5-二羧酸。

呋喃-2,5-二羧酸（FDCA）是可生物降解塑料的基础，可用于替代源自化石碳源的塑料。近年来，人们非常关注通过一系列的酶反应从生物基5-羟甲基糠醛（HMF）合成FDCA。芳醇氧化酶（AAO）和5-羟甲基糠醛氧化酶（HMFO）是葡萄糖-甲醇-胆碱类黄素酶，可用于通过三个连续的氧化反应从HMF生产FDCA，而无需辅助酶的辅助。这种具有挑战性的过程取决于原始醛基和所形成醛基的水合度，限速步骤在于中间体5-甲酰基-呋喃甲酸（FFCA）最终氧化为FDCA。尽管HMFO接受FFCA作为HMF反应途径的最终底物，AAO实际上无法将其氧化。在这里，我们设计了AAO，通过将其与HMFO进行结构排列和定向进化，将HMF逐步氧化为FDCA。随着HMF催化效率提高3倍，总转化率提高6倍，这种进化出的AAO为进一步工程化提供了一个有希望的起点，该工程旨在​​产生一种有效的生物催化剂，由HMF合成FDCA。