Department of Chemical, Biochemical and Environmental Engineering, University of Maryland, Baltimore County, Baltimore, MD, 21250, USA; College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, China.
Department of Chemical, Biochemical and Environmental Engineering, University of Maryland, Baltimore County, Baltimore, MD, 21250, USA; Department of Agronomy and Plant Breeding, University of Guilan, Rasht, Islamic Republic of Iran.
Metab Eng. 2019 Dec;56:60-68. doi: 10.1016/j.ymben.2019.08.017. Epub 2019 Aug 27.
Acetyl-CoA is the central metabolic node connecting glycolysis, Krebs cycle and fatty acids synthase. Plant-derived polyketides, are assembled from acetyl-CoA and malonyl-CoA, represent a large family of biological compounds with diversified bioactivity. Harnessing microbial bioconversion is considered as a feasible approach to large-scale production of polyketides from renewable feedstocks. Most of the current polyketide production platform relied on the lengthy glycolytic steps to provide acetyl-CoA, which inherently suffers from complex regulation with metabolically-costly cofactor/ATP requirements. Using the simplest polyketide triacetic acid lactone (TAL) as a testbed molecule, we demonstrate that acetate uptake pathway in oleaginous yeast (Yarrowia lipolytica) could function as an acetyl-CoA shortcut to achieve metabolic optimality in producing polyketides. We identified the metabolic bottlenecks to rewire acetate utilization for efficient TAL production in Y. lipolytica, including generation of the driving force for acetyl-CoA, malonyl-CoA and NADPH. The engineered strain, with the overexpression of endogenous acetyl-CoA carboxylase (ACC1), malic enzyme (MAE1) and a bacteria-derived cytosolic pyruvate dehydrogenase (PDH), affords robust TAL production with titer up to 4.76 g/L from industrial glacier acetic acid in shake flasks, representing 8.5-times improvement over the parental strain. The acetate-to-TAL conversion ratio (0.149 g/g) reaches 31.9% of the theoretical maximum yield. The carbon flux through this acetyl-CoA metabolic shortcut exceeds the carbon flux afforded by the native glycolytic pathways. Potentially, acetic acid could be manufactured in large-quantity at low-cost from Syngas fermentation or heterogenous catalysis (methanol carbonylation). This alternative carbon sources present a metabolic advantage over glucose to unleash intrinsic pathway limitations and achieve high carbon conversion efficiency and cost-efficiency. This work also highlights that low-cost acetic acid could be sustainably upgraded to high-value polyketides by oleaginous yeast species in an eco-friendly and cost-efficient manner.
乙酰辅酶 A 是连接糖酵解、三羧酸循环和脂肪酸合成酶的中心代谢节点。植物来源的聚酮化合物由乙酰辅酶 A 和丙二酰辅酶 A 组装而成，是一类具有多样化生物活性的生物化合物大家族。利用微生物生物转化被认为是从可再生原料大规模生产聚酮化合物的可行方法。目前大多数聚酮化合物生产平台都依赖于冗长的糖酵解步骤来提供乙酰辅酶 A，这本身就受到代谢成本高的辅因子/ATP 需求的复杂调节。我们以最简单的聚酮化合物三乙酸内酯 (TAL) 为测试分子，证明了油脂酵母（Yarrowia lipolytica）中的乙酸摄取途径可以作为乙酰辅酶 A 的捷径，以实现聚酮化合物生产中的代谢最优。我们确定了重新利用乙酸用于高效生产 TAL 的代谢瓶颈，包括为乙酰辅酶 A、丙二酰辅酶 A 和 NADPH 生成驱动力。通过过表达内源性乙酰辅酶 A 羧化酶 (ACC1)、苹果酸酶 (MAE1) 和细菌来源的细胞质丙酮酸脱氢酶 (PDH)，工程菌株能够从工业冰川乙酸中以 4.76 g/L 的摇瓶产量生产出 TAL，比亲本菌株提高了 8.5 倍。乙酸到 TAL 的转化率（0.149 g/g）达到理论最大产率的 31.9%。通过这条乙酰辅酶 A 代谢捷径的碳通量超过了天然糖酵解途径提供的碳通量。潜在地，可以通过合成气发酵或异相催化（甲醇羰基化）以低成本大量生产乙酸。与葡萄糖相比，这些替代碳源具有代谢优势，可以释放内在途径的限制，实现高碳转化率和成本效益。这项工作还强调，通过油脂酵母物种以环保和经济高效的方式，可以将低成本的乙酸可持续地升级为高价值的聚酮化合物。
