Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
University of Chinese Academy of Sciences, Beijing, China.
Biotechnol Bioeng. 2019 Feb;116(2):283-293. doi: 10.1002/bit.26827. Epub 2018 Nov 15.
One major challenge in the bioconversion of lignocelluloses into ethanol is to develop Saccharomyces cerevisiae strains that can utilize all available sugars in biomass hydrolysates, especially the d-xylose and l-arabinose that cannot be fermented by the S. cerevisiae strain naturally. Here, we integrated an l-arabinose utilization cassette (AUC) into the genome of an efficient d-xylose fermenting industrial diploid S. cerevisiae strain CIBTS0735 to make strain CIBTS1972. After evolving on arabinose, CIBTS1974 with excellent fermentation capacity was obtained. A comparison between genome sequences of strains CIBTS1974 and CIBTS1972 revealed that the copy number of the AUC had increased from 1 to 12. We then constructed the AUC null-mutant CIBTS1975 and gradually rescued the l-arabinose utilization defect by integrating AUC iteratively. On the other hand, the parental strain CIBTS0735 was able to acquire the same performance as CIBTS1974 by the direct introduction of 12 copies of the AUC; the performance was further improved by adding two more copies. Besides, we found that not the two transporters present in the AUC were both needed during l-arabinose utilization, GAL2 was necessary and STP2 was not essential. We have described for the first time that a high copy number of AUC is sufficient for the strain to metabolize l-arabinose efficiently independent of evolution.
在木质纤维素生物转化为乙醇的过程中，一个主要的挑战是开发能够利用生物质水解物中所有可用糖的酿酒酵母菌株，特别是不能被天然酿酒酵母菌株发酵的 d-木糖和 l-阿拉伯糖。在这里，我们将一个 l-阿拉伯糖利用盒（AUC）整合到高效 d-木糖发酵工业二倍体酿酒酵母菌株 CIBTS0735 的基因组中，得到菌株 CIBTS1972。在阿拉伯糖上进化后，获得了具有优异发酵能力的 CIBTS1974。菌株 CIBTS1974 和 CIBTS1972 的基因组序列比较表明，AUC 的拷贝数从 1 增加到 12。然后，我们构建了 AUC 缺失突变体 CIBTS1975，并通过迭代整合 AUC 逐渐恢复 l-阿拉伯糖利用缺陷。另一方面，亲本菌株 CIBTS0735 通过直接引入 12 个 AUC 拷贝能够获得与 CIBTS1974 相同的性能；通过再添加两个拷贝，性能得到进一步提高。此外，我们发现 l-阿拉伯糖利用过程中并不需要 AUC 中存在的两种转运蛋白，GAL2 是必需的，而 STP2 不是必需的。我们首次描述了高拷贝数的 AUC 足以使菌株在不依赖进化的情况下高效代谢 l-阿拉伯糖。
