DOE Joint BioEnergy Institute, 5885 Hollis St., Emeryville, CA 94608, United States; Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, United States.
DOE Joint BioEnergy Institute, 5885 Hollis St., Emeryville, CA 94608, United States; Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, United States; California Institute of Quantitative Biosciences (QB3), University of California, Berkeley, CA 94720, United States.
Metab Eng. 2018 Jul;48:52-62. doi: 10.1016/j.ymben.2018.05.018. Epub 2018 May 28.
Medium- and long-chain methyl ketones are fatty acid-derived compounds that can be used as biofuel blending agents, flavors and fragrances. However, their large-scale production from sustainable feedstocks is currently limited due to the lack of robust microbial biocatalysts. The oleaginous yeast Yarrowia lipolytica is a promising biorefinery platform strain for the production of methyl ketones from renewable lignocellulosic biomass due to its natively high flux towards fatty acid biosynthesis. In this study, we report the metabolic engineering of Y. lipolytica to produce long- and very long-chain methyl ketones. Truncation of peroxisomal β-oxidation by chromosomal deletion of pot1 resulted in the biosynthesis of saturated, mono-, and diunsaturated methyl ketones in the C-C range. Additional overexpression and peroxisomal targeting of a heterologous bacterial methyl ketone biosynthesis pathway yielded an initial titer of 151.5 mg/L of saturated methyl ketones. Dissolved oxygen concentrations in the cultures were found to substantially impact cell morphology and methyl ketone biosynthesis. Bioreactor cultivation under optimized conditions resulted in a titer of 314.8 mg/L of total methyl ketones, representing more than a 6000-fold increase over the parental strain. This work highlights the potential of Y. lipolytica to serve as chassis organism for the biosynthesis of acyl-thioester derived long- and very long-chain methyl ketones.
中链和长链甲基酮是脂肪酸衍生的化合物，可用作生物燃料混合剂、香料和香精。然而，由于缺乏强大的微生物生物催化剂，它们从可持续原料大规模生产目前受到限制。产油酵母 Yarrowia lipolytica 是一种很有前途的生物炼制平台菌株，可用于从可再生木质纤维素生物质生产甲基酮，因为其脂肪酸生物合成的固有高通量。在这项研究中，我们报告了 Y. lipolytica 的代谢工程，以生产长链和超长链甲基酮。通过染色体缺失 pot1 截断过氧化物酶体β-氧化导致 C-C 范围内的饱和、单不饱和和二不饱和甲基酮的生物合成。过表达和过氧化物酶体靶向异源细菌甲基酮生物合成途径进一步提高了初始产量，达到 151.5mg/L 的饱和甲基酮。发现培养物中的溶解氧浓度对细胞形态和甲基酮生物合成有很大影响。在优化条件下进行生物反应器培养，得到 314.8mg/L 的总甲基酮，比亲本菌株提高了 6000 多倍。这项工作突出了 Y. lipolytica 作为酰基辅酶 A 衍生长链和超长链甲基酮生物合成底盘生物的潜力。
