VTT Technical Research Centre of Finland Ltd., Solutions for Natural Resources and Environment, Tietotie 2, P.O. Box 1000, FI-02044 VTT, Espoo, Finland.
Appl Microbiol Biotechnol. 2017 Nov;101(22):8151-8163. doi: 10.1007/s00253-017-8547-3. Epub 2017 Oct 16.
The important platform chemicals ethylene glycol and glycolic acid were produced via the oxidative D-xylose pathway in the yeast Saccharomyces cerevisiae. The expression of genes encoding D-xylose dehydrogenase (XylB) and D-xylonate dehydratase (XylD) from Caulobacter crescentus and YagE or YjhH aldolase and aldehyde dehydrogenase AldA from Escherichia coli enabled glycolic acid production from D-xylose up to 150 mg/L. In strains expressing only xylB and xylD, 29 mg/L 2-keto-3-deoxyxylonic acid [(S)-4,5-dihydroxy-2-oxopentanoic acid] (2K3DXA) was produced and D-xylonic acid accumulated to ca. 9 g/L. A significant amount of D-xylonic acid (ca. 14%) was converted to 3-deoxypentonic acid (3DPA), and also, 3,4-dihydroxybutyric acid was formed. 2K3DXA was further converted to glycolaldehyde when genes encoding by either YagE or YjhH aldolase from E. coli were expressed. Reduction of glycolaldehyde to ethylene glycol by an endogenous aldo-keto reductase activity resulted further in accumulation of ethylene glycol of 14 mg/L. The possibility of simultaneous production of lactic and glycolic acids was evaluated by expression of gene encoding lactate dehydrogenase ldhL from Lactobacillus helveticus together with aldA. Interestingly, this increased the accumulation of glycolic acid to 1 g/L. The D-xylonate dehydratase activity in yeast was notably low, possibly due to inefficient Fe-S cluster synthesis in the yeast cytosol, and leading to D-xylonic acid accumulation. The dehydratase activity was significantly improved by targeting its expression to mitochondria or by altering the Fe-S cluster metabolism of the cells with FRA2 deletion.
通过酿酒酵母中的氧化 D-木糖途径，生产出重要的平台化学品乙二醇和乙醇酸。来自新月柄杆菌的 D-木糖脱氢酶（XylB）和 D-木酮酸脱水酶（XylD）以及大肠杆菌的 YagE 或 YjhH 醛缩酶和醛脱氢酶 AldA 的基因表达，使乙醇酸能够从 D-木糖生产到 150mg/L。在仅表达 xylB 和 xylD 的菌株中，产生了 29mg/L 的 2-酮-3-脱氧木酮酸[(S)-4,5-二羟基-2-氧戊酸]（2K3DXA），并且 D-木糖积累到约 9g/L。相当数量的 D-木糖（约 14%）转化为 3-脱氧戊糖（3DPA），同时也形成了 3,4-二羟基丁酸。当表达来自大肠杆菌的 YagE 或 YjhH 醛缩酶的基因时，2K3DXA 进一步转化为乙二醛。通过内源性醛酮还原酶活性将乙二醛还原为乙二醇，进一步导致 14mg/L 的乙二醇积累。通过表达乳杆菌的乳酸脱氢酶 ldhL 基因并与 aldA 一起表达，评估了同时生产乳酸和乙醇酸的可能性。有趣的是，这将乙醇酸的积累增加到 1g/L。酵母中的 D-木酮酸脱水酶活性明显较低，这可能是由于酵母细胞质中 Fe-S 簇合成效率低下，导致 D-木酮酸积累。通过将其表达靶向线粒体或通过 Fra2 缺失改变细胞的 Fe-S 簇代谢，可显著提高脱水酶的活性。
