Sugar phosphates cannot be produced easily by microbial fermentation because negatively-charged compounds cannot be secreted across intact cell membrane. D-xylulose 5-phosphate (Xu5P), a very expensive sugar phosphate, was synthesized from D-xylose and polyphosphate catalyzed by enzyme cascades in one pot. The synthetic enzymatic pathway comprised of xylose isomerase and xylulokinase was designed to produce Xu5P, along with a third enzyme, polyphosphate kinase, responsible for in site ATP regeneration. Due to the promiscuous activity of the ATP-based xylulokinase from a hyperthermophilic bacterium Thermotoga maritima on polyphosphate, the number of enzymes in the pathway was minimized to two without polyphosphate kinase. The reactions catalyzed by the two-enzyme and three-enzyme pathways were compared for Xu5P production, and the reaction conditions were optimized by examining effects of reaction temperature, enzyme ratio and substrate concentration. The optimized two-enzyme system produced 32 mM Xu5P from 50 mM xylose and polyphosphate after 36 h at 45°C. Biosynthesis of less costly Xu5P from D-xylose and polyphosphate could be highly feasible via this minimized two-enzyme pathway.

中文翻译：

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通过最小化的两个酶级联反应，由D-木糖和聚磷酸盐生物合成D-木酮糖5-磷酸。

由于微生物无法在完整的细胞膜上分泌带负电荷的化合物，因此无法通过微生物发酵轻松生产磷酸糖。D-木酮糖5-磷酸（Xu5P）是一种非常昂贵的糖磷酸酯，是由一锅中由酶级联反应催化的D-木糖和聚磷酸酯合成的。由木糖异构酶和木酮糖激酶组成的合成酶途径被设计为产生Xu5P，以及负责原位ATP再生的第三种酶多磷酸盐激酶。由于来自超嗜热细菌马氏热球菌的基于ATP的木酮糖酶对多磷酸盐的混杂活性，该途径中的酶数量被减至最少两种，而没有多磷酸盐激酶。比较了两种酶和三种酶途径催化的反应产生的Xu5P，通过考察反应温度，酶比和底物浓度的影响，优化了反应条件。优化的两种酶体系在45°C下放置36 h后，由50 mM木糖和多磷酸酯生成32 mM Xu5P。通过这种最小化的两种酶途径，由D-木糖和多磷酸盐生物合成成本较低的Xu5P可能是高度可行的。