Sayed Mahmoud, Ismail Mohamed, Sivasubramanian Anirudh, Kawano Riko, Li Chengsi, Glaser Sara Jonsdottir, Hatti-Kaul Rajni
Biotechnology and Applied Microbiology, Department of Process and Life Science Engineering, Kemicentrum, Lund University, Lund, SE-22100, Sweden.
Department of Botany and Microbiology, Faculty of Science, South Valley University, Qena, 83523, Egypt.
Microb Cell Fact. 2025 Mar 19;24(1):68. doi: 10.1186/s12934-025-02689-x.
2,5-Furandicarboxylic acid (FDCA) is a promising building block for biobased recyclable polymers and a platform for other potential biobased chemicals. The common route of its production is by oxidation of sugar-derived 5-hydroxymethylfurfural (HMF). Several reports on biocatalytic oxidation using whole microbial cells or enzymes have been reported, which offers potentially a greener alternative compared to the chemical process. HMF oxidases and aryl alcohol oxidases are the only enzymes able to catalyse the complete oxidation to FDCA, however at low concentrations and are subject to inhibition by the FFCA (5-formylfuran-2-carboxylic acid) intermediate. The present report presents a study on the oxidation of FFCA to FDCA using the obligately aerobic bacterium Gluconobacter oxydans and identification of the enzymes catalyzing the reaction.
Screening of three different strains showed G. oxydans DSM 50049 to possess the highest FFCA oxidation efficiency. Optimal reaction conditions for obtaining 100% conversion of 10 g/L (71 mM) FFCA to FDCA at 100% reaction yield were at pH 5, 30 °C and using 200 mg wwt /mL cells harvested at mild-exponential phase. In a reaction run at a 1 L scale using a total of 15 g/L (107 mM) FFCA supplied in a fed-batch mode, FDCA was obtained at a yield of 90% in 8.5 h. The product was recovered at 82% overall yield and 99% purity using a simple recovery process. Screening of several oxidoreductase enzymes from the gene sequences identified in the bacterial genome revealed two proteins annotated as membrane-bound aldehyde dehydrogenase (MALDH) and coniferyl aldehyde dehydrogenase (CALDH) to be the enzymes catalyzing the oxidization of FFCA.
The study shows G. oxydans DSM 50049 and its enzymes to be promising biocatalysts for use in the FDCA production process from biomass. The high reaction rate and yield motivate further studies on characterization of the identified enzymes exhibiting the FFCA oxidizing activity, which can be used to construct an enzyme cascade together e.g. with HMF oxidase or aryl alcohol oxidase for one-pot production of FDCA from 5-HMF.
2,5-呋喃二甲酸（FDCA）是一种用于生物基可回收聚合物的有前景的构建单元，也是其他潜在生物基化学品的一个平台。其常见的生产途径是通过糖衍生的5-羟甲基糠醛（HMF）的氧化。已经有几篇关于使用完整微生物细胞或酶进行生物催化氧化的报道，与化学过程相比，这可能提供一种更绿色的替代方法。HMF氧化酶和芳醇氧化酶是仅有的能够催化完全氧化为FDCA的酶，然而其浓度较低且会受到FFCA（5-甲酰基呋喃-2-羧酸）中间体的抑制。本报告介绍了一项关于使用专性需氧细菌氧化葡萄糖杆菌将FFCA氧化为FDCA的研究以及对催化该反应的酶的鉴定。
对三种不同菌株的筛选表明，氧化葡萄糖杆菌DSM 50049具有最高的FFCA氧化效率。在pH 5、30℃以及使用在温和指数期收获的200 mg湿重/ mL细胞的条件下，可实现10 g/L（71 mM）FFCA 100%转化为FDCA且反应产率达到100%。在1 L规模的反应中，采用分批补料模式总共供应15 g/L（107 mM）FFCA，在8.5小时内获得了产率为90%的FDCA。使用简单的回收工艺，产物的总回收率为82%，纯度为99%。从细菌基因组中鉴定的基因序列中筛选几种氧化还原酶，发现有两种蛋白质被注释为膜结合醛脱氢酶（MALDH）和松柏醛脱氢酶（CALDH）是催化FFCA氧化的酶。
该研究表明氧化葡萄糖杆菌DSM 50049及其酶是用于从生物质生产FDCA过程中的有前景的生物催化剂。高反应速率和产率促使进一步研究鉴定出的具有FFCA氧化活性的酶的特性，这些酶可用于构建酶级联反应，例如与HMF氧化酶或芳醇氧化酶一起用于从5-HMF一锅法生产FDCA。
