Microbial catalysis has recently emerged as one of the most promising approaches in oligosaccharide synthesis. However, despite significant progress, microbial synthesis still requires much improvement in efficiency and in reduction of process complexity. Additionally, given the stunning diversity and many varied applications of glycans, broadening the range of glycans accessible via microbial synthesis is of paramount importance. Major challenges in microbial synthesis include catabolite repression and high cellular energy requirement. Here we demonstrated a new approach to overcome these challenges by directly tapping into the cellular “power house,” the TCA cycle, to provide the cellular energy for synthesis. This approach not only circumvents catabolite repression but also eliminates acidic glycolysis by-products. As such, the whole-cell biocatalysis can be carried out without sophisticated fed-batch feeding and pH control in the synthesis stage. The system could achieve several grams per liter (3–4 g/L) within a 24-h period in shaker flask cultivation for two targets, fucosyllactose and fucosyllactulose, demonstrating efficiency of the biocatalyst developed and its applicability to both natural and non-natural targets. To the best of our knowledge, this is the first use of TCA cycle intermediates as the energy source for oligosaccharide synthesis and the first description of successful synthesis of fucosyllactulose with titers in several grams per liter.

中文翻译：

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TCA循环驱动岩藻糖基化低聚糖的合成

微生物催化最近已成为寡糖合成中最有希望的方法之一。然而，尽管取得了重大进展，但微生物合成仍需要在效率和降低工艺复杂性方面进行很大的改进。另外，考虑到聚糖的惊人多样性和许多不同的应用，扩大通过微生物合成可获得的聚糖的范围是最重要的。微生物合成中的主要挑战包括分解代谢物抑制和高细胞能量需求。在这里，我们展示了一种通过直接利用细胞“动力之家” TCA循环来克服这些挑战的新方法，为合成提供细胞能量。这种方法不仅可以避免分解代谢物的抑制，而且可以消除酸性糖酵解副产物。因此，在合成阶段，无需复杂的补料分批进料和pH值控制即可进行全细胞生物催化。在摇瓶培养中，岩藻糖半乳糖和岩藻糖半乳糖这两个目标可在24小时内达到每升几克（3-4 g / L），这证明了所开发生物催化剂的效率及其对天然和非天然生物的适用性目标。据我们所知，这是首次将TCA循环中间体用作寡糖合成的能源，并且首次成功地成功合成了岩藻糖基果糖，其滴度为每升几克。在摇瓶培养中，岩藻糖半乳糖和岩藻糖半乳糖这两个目标可在24小时内达到每升几克（3-4 g / L），这证明了所开发生物催化剂的效率及其对天然和非天然生物的适用性目标。据我们所知，这是首次将TCA循环中间体用作寡糖合成的能源，并且首次成功地成功合成了岩藻糖基果糖，其滴度为每升几克。在摇瓶培养中，岩藻糖半乳糖和岩藻糖半乳糖这两个目标可在24小时内达到每升几克（3-4 g / L），这证明了所开发生物催化剂的效率及其对天然和非天然生物的适用性目标。据我们所知，这是首次将TCA循环中间体用作寡糖合成的能源，并且首次成功地成功合成了岩藻糖基果糖，其滴度为每升几克。