Department of Life Sciences and Chemistry, Jacobs University Bremen gGmbH, Campus Ring 1, 28759 Bremen, Germany.
School of Biomedical and Healthcare Sciences, Plymouth University Peninsula Schools of Medicine and Dentistry, Plymouth, Devon, PL4 8AA, UK.
FEMS Yeast Res. 2018 May 1;18(3). doi: 10.1093/femsyr/foy019.
Glycerol offers several advantages as a substrate for biotechnological applications. An important step toward using the popular production host Saccharomyces cerevisiae for glycerol-based bioprocesses has been the fact that in recent studies commonly used S. cerevisiae strains were engineered to grow in synthetic medium containing glycerol as the sole carbon source. For metabolic engineering projects of S. cerevisiae growing on glycerol, characterized promoters are missing. In the current study, we used transcriptome analysis and a yECitrine-based fluorescence reporter assay to select and characterize 25 useful promoters. The promoters of the genes ALD4 and ADH2 showed 4.2-fold and 3-fold higher activities compared to the well-known strong TEF1 promoter. Moreover, the collection contains promoters with graded activities in synthetic glycerol medium and different degrees of glucose repression. To demonstrate the general applicability of the promoter collection, we successfully used a subset of the characterized promoters with graded activities in order to optimize growth on glycerol in an engineered derivative of CEN.PK, in which glycerol catabolism exclusively occurs via a non-native DHA pathway.
甘油作为生物技术应用的基质具有几个优势。将广受欢迎的生产宿主酿酒酵母用于基于甘油的生物工艺的重要一步是，在最近的研究中，通常使用的酿酒酵母菌株被设计为在含有甘油作为唯一碳源的合成培养基中生长。对于在甘油上生长的酿酒酵母的代谢工程项目，缺少特征性启动子。在本研究中，我们使用转录组分析和基于 yECitrine 的荧光报告测定法来选择和表征 25 个有用的启动子。与知名的强 TEF1 启动子相比，ALD4 和 ADH2 基因的启动子的活性分别高出 4.2 倍和 3 倍。此外，该集合包含在合成甘油培养基中具有分级活性的启动子，以及不同程度的葡萄糖抑制。为了证明启动子集合的通用性，我们成功地使用了具有分级活性的特征启动子子集，以便在 CEN.PK 的工程衍生物中优化甘油的生长，其中甘油代谢仅通过非天然 DHA 途径发生。
