College of Chemical Engineering, Fujian Engineering Research Center of Advanced Manufacturing Technology for Fine Chemicals, Fuzhou University, Fuzhou 350108, China.
College of Chemical Engineering, Fujian Engineering Research Center of Advanced Manufacturing Technology for Fine Chemicals, Fuzhou University, Fuzhou 350108, China; Qingyuan Innovation Laboratory, Quanzhou 362801, China.
J Biotechnol. 2024 Nov 10;394:103-111. doi: 10.1016/j.jbiotec.2024.08.012. Epub 2024 Aug 23.
D-allulose, a naturally occurring monosaccharide, is present in small quantities in nature. It is considered a valuable low-calorie sweetener due to its low absorption in the digestive tract and zero energy for growth. Most of the recent efforts to produce D-allulose have focused on in vitro enzyme catalysis. However, microbial fermentation is emerging as a promising alternative that offers the advantage of combining enzyme manufacturing and product synthesis within a single bioreactor. Here, a novel approach was proposed for the efficient biosynthesis of D-allulose from glycerol using metabolically engineered Escherichia coli. FbaA, Fbp, AlsE, and A6PP were used to construct the D-allulose synthesis pathway. Subsequently, PfkA, PfkB, and Pgi were disrupted to block the entry of the intermediate fructose-6-phosphate (F6P) into the Embden-Meyerhof-Parnas (EMP) and pentose phosphate (PP) pathways. Additionally, GalE and FryA were inactivated to reduce D-allulose consumption by the cells. Finally, a fed-batch fermentation process was implemented to optimize the performance of the cell factory. As a result, the titer of D-allulose reached 7.02 g/L with a maximum yield of 0.287 g/g.
D-阿洛酮糖是一种天然存在的单糖，在自然界中含量很少。由于其在消化道中的吸收量低，并且对生长没有能量贡献，因此被认为是一种有价值的低卡路里甜味剂。最近大多数生产 D-阿洛酮糖的努力都集中在体外酶催化上。然而，微生物发酵作为一种很有前途的替代方法正在出现，它具有将酶制造和产品合成在单个生物反应器中结合的优势。在这里，提出了一种使用代谢工程大肠杆菌从甘油高效生物合成 D-阿洛酮糖的新方法。使用 FbaA、Fbp、AlsE 和 A6PP 构建 D-阿洛酮糖合成途径。随后，破坏 PfkA、PfkB 和 Pgi 以阻止中间产物果糖-6-磷酸（F6P）进入 EMP 和磷酸戊糖（PP）途径。此外，失活 GalE 和 FryA 以减少细胞对 D-阿洛酮糖的消耗。最后，实施分批补料发酵过程以优化细胞工厂的性能。结果，D-阿洛酮糖的浓度达到 7.02 g/L，最大得率为 0.287 g/g。
