Key Laboratory of Fermentation Engineering (Ministry of Education), Hubei Provincial Key Laboratory of Industrial Microbiology, College of Bioengineering, Hubei University of Technology, Wuhan 430068, China.
Microb Cell Fact. 2013 May 2;12:41. doi: 10.1186/1475-2859-12-41.
Current methods of ethanol production from lignocelluloses generate a mixture of sugars, primarily glucose and xylose; the fermentation cells are always exposed to stresses like high temperature and low nutritional conditions that affect their growth and productivity. Stress-tolerant strains capable of using both glucose and xylose to produce ethanol with high yield are highly desirable.
A recombinant Zymomonas mobilis (Z. mobilis) designated as HYMX was constructed by integrating seven genes (Pfu-sHSP, yfdZ, metB, xylA, xylB, tktA and talB) into the genome of Z. mobilis CP4 (CP4) via Tn5 transposon in the present study. The small heat shock protein gene (Pfu-sHSP) from Pyrococcus furious (P. furious) was used to increase the heat-tolerance, the yfdZ and metB genes from E. coli were used to decrease the nutritional requirement. To overcome the bottleneck of CP4 being unable to use pentose, xylose catabolic genes (xylA, xylB, tktA and talB) from E. coli were integrated into CP4 also for construction of the xylose utilizing metabolic pathway.
The genomic integration confers on Z. mobilis the ability to grow in medium containing xylose as the only carbon source, and to grow in simple chemical defined medium without addition of amino acid. The HYMX demonstrated not only the high tolerance to unfavorable stresses like high temperature and low nutrient, but also the capability of converting both glucose and xylose to ethanol with high yield at high temperature. What's more, these genetic characteristics were stable up to 100 generations on nonselective medium. Although significant improvements were achieved, yeast extract is needed for ethanol production.
目前从木质纤维素生产乙醇的方法会生成糖的混合物，主要是葡萄糖和木糖；发酵细胞总是会受到高温和低营养条件等压力的影响，这会影响它们的生长和生产力。能够利用葡萄糖和木糖高产生产乙醇的耐应激菌株是非常理想的。
本研究通过 Tn5 转座子将七个基因（Pfu-sHSP、yfdZ、metB、xylA、xylB、tktA 和 talB）整合到 Zymomonas mobilis CP4（CP4）的基因组中，构建了一株名为 HYMX 的重组 Zymomonas mobilis（Z. mobilis）。来自 Pyrococcus furious（P. furious）的小热休克蛋白基因（Pfu-sHSP）用于提高耐热性，来自大肠杆菌的 yfdZ 和 metB 基因用于减少营养需求。为了克服 CP4 无法利用戊糖的瓶颈，还将大肠杆菌的木糖分解代谢基因（xylA、xylB、tktA 和 talB）整合到 CP4 中，构建了木糖利用代谢途径。
基因组整合使 Z. mobilis 能够在仅含有木糖作为碳源的培养基中生长，并能够在不添加氨基酸的简单化学定义培养基中生长。HYMX 不仅表现出对高温和低营养等不利压力的高耐受性，而且还能够在高温下将葡萄糖和木糖高效转化为乙醇。此外，这些遗传特性在非选择性培养基上稳定保持了 100 代。尽管取得了显著的进展，但仍需要酵母提取物来生产乙醇。
