Beijing Advanced Innovation Center for Soft Matter Science and Engineering, College of Life Science and Technology, Beijing University of Chemical Technology, No.15 North Third Ring Road East, Chaoyang District, Beijing, 100029, People's Republic of China.
Department of Biology and Biological Engineering, Chalmers University of Technology, Gothenburg, Sweden.
Microb Cell Fact. 2020 Dec 10;19(1):226. doi: 10.1186/s12934-020-01493-z.
Saccharomyces cerevisiae is being exploited as a cell factory to produce fatty acids and their derivatives as biofuels. Previous studies found that both precursor supply and fatty acid metabolism deregulation are essential for enhanced fatty acid synthesis. A bacterial pyruvate dehydrogenase (PDH) complex expressed in the yeast cytosol was reported to enable production of cytosolic acetyl-CoA with lower energy cost and no toxic intermediate.
Overexpression of the PDH complex significantly increased cell growth, ethanol consumption and reduced glycerol accumulation. Furthermore, to optimize the redox imbalance in production of fatty acids from glucose, two endogenous NAD-dependent glycerol-3-phosphate dehydrogenases were deleted, and a heterologous NADP-dependent glyceraldehyde-3-phosphate dehydrogenase was introduced. The best fatty acid producing strain PDH7 with engineering of precursor and co-factor metabolism could produce 840.5 mg/L free fatty acids (FFAs) in shake flask, which was 83.2% higher than the control strain YJZ08. Profile analysis of free fatty acid suggested the cytosolic PDH complex mainly resulted in the increases of unsaturated fatty acids (C16:1 and C18:1).
We demonstrated that cytosolic PDH pathway enabled more efficient acetyl-CoA provision with the lower ATP cost, and improved FFA production. Together with engineering of the redox factor rebalance, the cytosolic PDH pathway could achieve high level of FFA production at similar levels of other best acetyl-CoA producing pathways.
酿酒酵母被用作细胞工厂来生产脂肪酸及其衍生物作为生物燃料。先前的研究发现，前体供应和脂肪酸代谢失调对于增强脂肪酸合成都是必不可少的。据报道，在酵母细胞质中表达的细菌丙酮酸脱氢酶（PDH）复合物能够以更低的能量成本和没有有毒中间体产生细胞质乙酰辅酶 A。
PDH 复合物的过表达显著增加了细胞生长、乙醇消耗和减少了甘油积累。此外，为了优化从葡萄糖生产脂肪酸过程中的氧化还原失衡，删除了两个内源性 NAD 依赖性甘油-3-磷酸脱氢酶，并引入了一个异源 NADP 依赖性甘油醛-3-磷酸脱氢酶。通过对前体和辅助因子代谢进行工程改造的最佳脂肪酸生产菌株 PDH7 可以在摇瓶中产生 840.5mg/L 的游离脂肪酸（FFA），比对照菌株 YJZ08 高 83.2%。游离脂肪酸的谱分析表明，细胞质 PDH 复合物主要导致不饱和脂肪酸（C16:1 和 C18:1）增加。
我们证明了细胞质 PDH 途径能够以更低的 ATP 成本提供更有效的乙酰辅酶 A，并提高了 FFA 的产量。与氧化还原因子再平衡的工程改造相结合，细胞质 PDH 途径可以在类似的其他最佳乙酰辅酶 A 产生途径的水平上实现高水平的 FFA 生产。
