ORP敏感型元件增强酿酒酵母对木质纤维素水解液混合抑制物的耐受性

纤维素乙醇生产中原料预处理过程产生的抑制物严重影响酵母细胞活性，导致乙醇发酵速率减慢，残糖浓度升高，乙醇对原料的表观收率降低。提高细胞对单一抑制物耐受性的研究已大量开展，但构建耐受性菌种应对真实水解液中混合抑制物还具有难度。本项目拟针对胞内氧化还原电位（ORP）与胁迫耐受性的关联，探索解决该问题的途径。首先,根据细胞受抑制时胞内ORP的改变，利用在微生物细胞中已经鉴定的ORP敏感型转录系统,设计能针对性响应ROS及还原力等的敏感型元件。其次，使用多个ORP敏感型元件，精确响应胞内ROS增高、还原力下降及能量损耗等状态，开启下游特定蛋白表达，智能地调控细胞的胁迫响应状态。最后，辅助胞外过程工程的控制策略强化细胞内元件功能效果。预期取得的研究进展,不仅可以加深对ORP提高酵母细胞胁迫耐受性的科学认识，还可以为构建耐受混合抑制物菌株奠定基础，解决纤维素乙醇发酵速率慢和乙醇表观收率低的突出问题。

The toxic by-products released during the pretreatment of lignocellulosic biomass inhibit yeast viability, which consequently prolongs ethanol fermentation time with more sugars remained unfermented (stuck fermentation) and substantially compromises ethanol yield that is calculated based on the total sugars feeding into the fermentation system without deduction of residual sugars. Tolerance of Saccharomyces cerevisiae to individual inhibitors has been intensively studied. However, it is still challenging for S. cerevisiae to tolerate lignocellulose hydrolysate which contains various inhibitors with complicated synergistic effect. In this project, molecular mechanism underlying the correlation between intracellular redox potential (ORP) and the stress tolerance of S. cerevisiae will be elucidated. Firstly, based on the ORP response under stressful conditions, ORP sensing elements will be constructed based on natural redox regulatory transcription systems in either prokaryotic or eukaryotic microorganisms, which are able to recognize specific ORP mediators such as ROS, NAD(P)H, oxygen and ATP. Secondly, the target genes will be induced precisely by ROS accumulation, depletion of reducing power, and ATP shortage through each corresponding element or their combination. Finally, the assistance of environmental ORP regulation will be investigated to enhance the function of ORP sensing elements through bioprocess engineering strategies to improve ethanol production from lignocellulosic biomass. The progress will enrich scientific understanding of yeast stress tolerance under ORP control conditions, and in the meantime benefit the construction of robust strains that tolerate the inhibitory effect of inhibitors in the hydrolysate to produce ethanol from lignocellulosic biomass more efficiently.