Abstract D-tagatose (D-tag) is a natural yet rare hexose that can be used as a functional sweetener owing to its hypoglycemic and anticaries properties. A four-step enzymatic cascade reaction was performed to synthesize d -tagatose from maltodextrin. In this study, we constructed a catalytic cascade system by co-immobilizing α-glucan phosphorylase (αGP) and phosphoglucomutase (PGM) on Duolite A568 exchange resin to produce glucose 6-phosphate (G6P), a precursor of d -tagatose. Our results showed that the thermal stability of co-immobilized αGP&PGM was enhanced comparing to free enzymes. Co-immobilized αGP&PGM retained 63.4% activity after incubating at 75 °C for 36 h, whereas free enzymes retained only 12% activity. Moreover, the relative activity of co-immobilized αGP&PGM was consistently maintained above 78.5% after eight cycles of reuse, and its storage stability was approximately eleven times higher than that of the free enzymes after storage at 4 °C for 60 days. Notably, we successfully co-immobilized dual enzymes on the resin using poly (ethylene glycol) diglycidyl ether, which was found to be more effective than glutaraldehyde. The results suggested that co-immobilized αGP&PGM could act as a promising catalyst for the one-pot production of G6P and be used in the starting pathway for the further production of d -tagatose.

中文翻译：

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双酶共固定化用于从麦芽糊精中一锅法生产葡萄糖 6-磷酸

摘要 D-塔格糖（D-tag）是一种天然且稀有的己糖，由于其具有降血糖和抗龋齿的特性，可用作功能性甜味剂。进行四步酶促级联反应以从麦芽糖糊精合成 d-塔格糖。在这项研究中，我们通过将 α-葡聚糖磷酸化酶 (αGP) 和磷酸葡糖变位酶 (PGM) 共同固定在 Duolite A568 交换树脂上以产生 6-磷酸葡萄糖 (G6P)，即 d-塔格糖的前体，构建了一个催化级联系统。我们的结果表明，与游离酶相比，共固定化的 αGP&PGM 的热稳定性得到增强。共固定化的 αGP&PGM 在 75°C 孵育 36 小时后保留了 63.4% 的活性，而游离酶仅保留了 12% 的活性。此外，共固定化的 αGP&PGM 的相对活性始终保持在 78 以上。8 次重复使用后 5%，在 4°C 下储存 60 天后，其储存稳定性比游离酶高约 11 倍。值得注意的是，我们使用聚（乙二醇）二缩水甘油醚成功地将双酶共固定在树脂上，发现它比戊二醛更有效。结果表明，共固定化的 αGP&PGM 可以作为一锅法生产 G6P 的有希望的催化剂，并用于进一步生产 d-塔格糖的起始途径。