Anaerobic xylose fermentation by recombinant Saccharomyces cerevisiae carrying XYL1, XYL2, and XKS1 in mineral medium chemostat cultures
Anaerobic xylose fermentation by recombinant Saccharomyces cerevisiae carrying XYL1, XYL2, and XKS1 in mineral medium chemostat cultures
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DOI：
10.1128/aem.66.8.3381-3386.2000
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发表时间：
2000-08-01
影响因子：
4.4
通讯作者：
Hahn-Hägerdal, B
中科院分区：
文献类型：
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作者：
Eliasson, A;Christensson, C;Hahn-Hägerdal, B
对于从木质纤维素生产乙醇，木糖发酵是经济上的必需，酿酒酵母已通过木糖利用途径进行代谢工程改造。然而，尚未实现用葡萄糖实现的高乙醇产率和生产率。为了定量分析重组酿酒酵母在木糖-葡萄糖混合物代谢过程中的代谢通量，我们构建了稳定的木糖利用重组菌株TMB 3001，来自树干毕赤酵母的XYL1和XYL2基因分别编码木糖还原酶（XR）和木糖醇脱氢酶（XDH），并 编码木酮糖激酶 (XK) 的内源 XKS1 基因在 PGK1 启动子的控制下被整合到酿酒酵母 CEN.PK 113-7A 的染色体 HIS3 基因座中。该菌株的XR、XDH和XK活性分别为0.4至0.5、2.7至3.4和1.5至1.7 U/mg，并且在连续发酵中稳定超过40代。重组酿酒酵母从木糖形成厌氧乙醇的过程尚属首次。然而，该菌株仅在有氧存在的情况下在木糖上生长。从木糖-葡萄糖混合物中获得 0.45 至 0.50 mmol C/mmol C (0.35 至 0.38 gig) 的乙醇产量和 9.7 至 13.2 mmol C h(-1) g (干重)细胞(-1) (0.24 至 0.30 g h(-1) g [干重]细胞(-1))。厌氧恒化培养物， 稀释率为0.06 h(-1)。假设葡萄糖上的乙醇产量恒定，则木糖上的厌氧乙醇产量估计为0.27molC/(木糖的molC1)(0.21g)。木糖摄取率随着饲料中木糖浓度的增加而增加，从饲料中木糖与葡萄糖比例为1:3时细胞(-1)的3.3 mmol C h(-1) g(干重)到饲料比例为3:1时细胞(-1)的6.8 mmol C h(-1) g(干重)，饲料含量为木糖/升15 g和5 g的 葡萄糖/升，木糖通量比葡萄糖通量低2.2倍，表明运输限制了木糖通量。
For ethanol production from lignocellulose, the fermentation of xylose is an economic necessity, Saccharomyces cerevisiae has been metabolically engineered with a xylose-utilizing pathway. However, the high ethanol yield and productivity seen with glucose have not yet been achieved. To quantitatively analyze metabolic fluxes in recombinant S. cerevisiae during metabolism of xylose-glucose mixtures, we constructed a stable xylose-utilizing recombinant strain, TMB 3001, The XYL1 and XYL2 genes from Pichia stipitis, encoding xylose reductase (XR) and xylitol dehydrogenase (XDH), respectively, and the endogenous XKS1 gene, encoding xylulokinase (XK), under control of the PGK1 promoter were integrated into the chromosomal HIS3 locus of S. cerevisiae CEN.PK 113-7A. The strain expressed XR, XDH, and XK activities of 0.4 to 0.5, 2.7 to 3.4, and 1.5 to 1.7 U/mg, respectively, and was stable for more than 40 generations in continuous fermentations. Anaerobic ethanol formation from xylose by recombinant S. cerevisiae was demonstrated far the first time. However, the strain grew on xylose only in the presence of oxygen. Ethanol yields of 0.45 to 0.50 mmol of C/mmol of C (0.35 to 0.38 gig) and productivities of 9.7 to 13.2 mmol of C h(-1) g (dry weight) of cells(-1) (0.24 to 0.30 g h(-1) g [dry weight] of cells(-1)) were obtained from xylose-glucose mixtures in anaerobic chemostat cultures, with a dilution rate of 0.06 h(-1). The anaerobic ethanol yield on xylose was estimated at 0.27 mol of C/(mol of C of xylose) (0.21 gig), assuming a constant ethanol yield on glucose. The xylose uptake rate increased with increasing xylose concentration in the feed, from 3.3 mmol of C h(-1) g (dry weight) of cells(-1) when the xylose-to-glucose ratio in the feed was 1:3 to 6.8 mmol of C h(-1) g (dry weight) of cells(-1) when the feed ratio was 3:1, With a feed content of 15 g of xylose/liter and 5 g of glucose/liter, the xylose flux was 2.2 times lower than the glucose flux, indicating that transport limits the xylose flux.