Wang Zedi, Zhou Linlin, Lu Minrui, Zhang Yuwei, Perveen Samina, Zhou Huarong, Wen Zhiqiang, Xu Zhaoxian, Jin Mingjie
School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing, 210094, China.
Appl Microbiol Biotechnol. 2021 Feb;105(4):1745-1758. doi: 10.1007/s00253-021-11130-3. Epub 2021 Feb 1.
Yarrowia lipolytica strain is a promising cell factory for the conversion of lignocellulose to biofuels and bioproducts. Despite the inherent robustness of this strain, further improvements to lignocellulose-derived inhibitors toxicity tolerance of Y. lipolytica are also required to achieve industrial application. Here, adaptive laboratory evolution was employed with increasing concentrations of ferulic acid. The adaptive laboratory evolution experiments led to evolve Y. lipolytica strain yl-XYL + FA4 with increased tolerance to ferulic acid as compared to the parental strain. Specifically, the evolved strain could tolerate 1.5 g/L ferulic acid, whereas 0.5 g/L ferulic acid could cause about 90% lethality of the parental strain. Transcriptome analysis of the evolved strain revealed several targets underlying toxicity tolerance enhancements. YALI0_E25201g, YALI0_F05984g, YALI0_B18854g, and YALI0_F16731g were among the highest upregulated genes, and the beneficial contributions of these genes were verified via reverse engineering. Recombinant strains with overexpressing each of these four genes obtained enhanced tolerance to ferulic acid as compared to the control strain. Fortunately, recombinant strains with overexpression of YALI0_E25201g, YALI0_B18854g, and YALI0_F16731g individually also obtained enhanced tolerance to vanillic acid. Overall, this work demonstrated a whole strain improvement cycle by "non-rational" metabolic engineering and presented new targets to modify Y. lipolytica for microbial lignocellulose valorization. KEY POINTS: • Adaptive evolution improved the ferulic acid tolerance of Yarrowia lipolytica • Transcriptome sequence was applied to analyze the ferulic acid tolerate strain • Three genes were demonstrated for both ferulic acid and vanillic acid tolerance.
解脂耶氏酵母菌株是将木质纤维素转化为生物燃料和生物产品的一种很有前景的细胞工厂。尽管该菌株具有内在的稳健性，但为了实现工业化应用，还需要进一步提高解脂耶氏酵母对木质纤维素衍生抑制剂毒性的耐受性。在此，采用适应性实验室进化方法，逐步增加阿魏酸的浓度。适应性实验室进化实验培育出了解脂耶氏酵母菌株yl-XYL + FA4，与亲本菌株相比，其对阿魏酸的耐受性增强。具体而言，进化后的菌株能够耐受1.5 g/L的阿魏酸，而0.5 g/L的阿魏酸会导致亲本菌株约90%的致死率。对进化菌株的转录组分析揭示了几个耐受性增强的潜在靶点。YALI0_E25201g、YALI0_F05984g、YALI0_B18854g和YALI0_F16731g是上调程度最高的基因之一，通过逆向工程验证了这些基因的有益作用。与对照菌株相比，分别过表达这四个基因的重组菌株对阿魏酸的耐受性增强。幸运的是，单独过表达YALI0_E25201g、YALI0_B18854g和YALI0_F16731g的重组菌株对香草酸的耐受性也增强。总体而言，这项工作通过“非理性 ”代谢工程展示了一个完整的菌株改良循环，并为改造解脂耶氏酵母以实现微生物木质纤维素增值提出了新的靶点。要点：• 适应性进化提高了解脂耶氏酵母对阿魏酸的耐受性 • 应用转录组序列分析阿魏酸耐受菌株 • 三个基因被证明对阿魏酸和香草酸均具有耐受性
