Department of Environmental Science and Engineering, College of Architecture and Environment, Sichuan University, No. 24, South Section 1, First Ring Road, Chengdu, Sichuan, 610065, China.
College of Pharmacy and and Biological Engineering, Chengdu University, No. 1763, Chengluo Road, Chengdu, 610106, China.
Appl Microbiol Biotechnol. 2017 Oct;101(20):7741-7753. doi: 10.1007/s00253-017-8494-z. Epub 2017 Sep 13.
It is of utmost importance to construct industrial xylose-fermenting Saccharomyces cerevisiae strains for lignocellulosic bioethanol production. In this study, two xylose isomerase-based industrial S. cerevisiae strains, O7 and P5, were constructed by δ-integration of the xylose isomerase (XI) gene xylA from the fungus Orpinomyces sp. and from the bacterium Prevotella ruminicola, respectively. The xylose consumption of the strains O7 and P5 at 48-h fermentation was 17.71 and 26.10 g/L, respectively, in synthetic medium with xylose as the sole sugar source. Adaptive evolution further improved the xylose fermentation capacity of the two strains to 51.0 and 28.9% in average, respectively. The transcriptomes of these two strains before and after evolution were analyzed using RNA-Seq. The expression levels of the genes involved in cell integrity, non-optimal sugar utilization, and stress response to environment were significantly up-regulated after evolution and did not depend on the origin of xylA; the expression levels of the genes involved in transmembrane transport, rRNA processing, cytoplasmic translation, and other processes were down-regulated. The expression of genes involved in central carbon metabolism was fine-tuned after the evolution. The analysis of transcription factors (TFs) indicated that most of the genes with significant differential expression were regulated by the TFs related to cell division, DNA damage response, or non-optimal carbon source utilization. The results of this study could provide valuable references for the construction of efficient xylose-fermenting XI strains.
构建用于木质纤维素生物乙醇生产的工业木糖发酵酿酒酵母菌株至关重要。在本研究中,通过δ-整合木糖异构酶(XI)基因 xylA,分别从真菌 Orpinomyces sp. 和细菌 Prevotella ruminicola 构建了两种基于木糖异构酶的工业酿酒酵母菌株 O7 和 P5。在以木糖为唯一糖源的合成培养基中,菌株 O7 和 P5 在 48 小时发酵时的木糖消耗分别为 17.71 和 26.10 g/L。适应性进化进一步将这两个菌株的木糖发酵能力提高到平均 51.0%和 28.9%。使用 RNA-Seq 分析了这两个菌株在进化前后的转录组。细胞完整性、非最佳糖利用以及对环境应激的反应相关基因的表达水平在进化后显著上调,并且不依赖于 xylA 的起源;与跨膜运输、rRNA 加工、细胞质翻译和其他过程相关的基因的表达水平下调。中央碳代谢相关基因的表达在进化后得到了精细调控。转录因子(TFs)的分析表明,大多数差异表达基因受到与细胞分裂、DNA 损伤反应或非最佳碳源利用相关的 TF 调控。本研究的结果可为构建高效木糖发酵 XI 菌株提供有价值的参考。