Construction of a Synthetic Metabolic Pathway for the Carbon-Conserving Biosynthesis of Value-Added Products from Ethylene Glycol and Glycolaldehyde

乙二醇和乙醇醛增值产品节碳生物合成的合成代谢途径的构建

• 批准号: 413016763
• 负责人: Professor Dr.-Ing. Thomas Walther
• 金额: --
• 依托单位: Institut für Naturstofftechnik
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2018
• 资助国家: 德国
• 起止时间: 2017-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/413016763?language=en
• 关键词: Construction; Synthetic; Metabolic; Pathway; Carbon

Development of the bio-based economy requires the exploitation of new carbon sources to satisfy the increasing demand for organic substrates that can be used in microbial product syntheses. The most sustainable strategy to meet this demand is to use carbon dioxide (CO2) as renewable carbon source for production of chemical compounds. To make CO2 accessible as a substrate for heterotrophic microorganisms, it is first chemically converted to methanol or synthesis gas. Both compounds are attractive carbon sources from which the whole panel of industrially relevant chemicals can be produced by microorganisms. However, the performance of these microbial product syntheses is limited by the extremely high oxygen demand of methanol-based biosyntheses and comparatively slow phase transition of the gaseous substrates synthesis gas and oxygen into the liquid culture medium. To solve these technological problems, the Institute of Natural Materials Technology of the Technische Universität Dresden proposes to chemically convert synthesis gas and methanol into the liquid and non-volatile C2-compounds ethylene glycol (EG) and glycolaldehyde (GA). Using these liquid compounds as carbon sources solves the phase transition problem. In addition, conversion of these molecules into value-added products (by the herein proposed synthetic metabolic pathways) has a significantly lower oxygen demand than when directly using methanol as the substrate.Various bacteria are capable of using EG and GA as a carbon source. However, natural EG-assimilating pathways contain several CO2-releasing reactions, which makes them particularly unsuitable for acetyl-CoA dependent product syntheses (maximum carbon yield of acetyl-CoA on EG is 50 %). In this research project, a synthetic metabolic pathway is proposed which enables carbon-conserving biosynthesis of acetyl-CoA from EG. The pathway relies on the initial oxidation of EG to GA and the aldol addition of GA to glyceraldehyde 3-phosphate (GAP). The reaction product arabinose 5-phosphate is further processed by several reaction steps to yield acetyl-CoA and to regenerate the GA-acceptor GAP. The function of the synthetic pathway will be first demonstrated in a cell-free reaction system by using isolated enzymes. This work is followed by implementation of the pathway in an engineered Escherichia coli strain. In vivo function of the pathway will be demonstrated by showing biosynthesis of mevalonate which is derived from acetyl-CoA. Optimization of EG-assimilation and mevalonate production will be guided by systems level physiological analyses.Our study makes an important contribution to the exploitation of alternative carbon sources for microbial product syntheses.

生物经济的发展要求开发新的碳源，以满足对可用于微生物产品合成的有机底物日益增长的需求。满足这一需求的最可持续的战略是使用二氧化碳(CO2)作为可再生碳源来生产化合物。为了使二氧化碳成为异养微生物的底物，首先将其化学转化为甲醇或合成气。这两种化合物都是有吸引力的碳源，微生物可以利用这些碳源生产所有与工业相关的化学品。然而，这些微生物产物合成的性能受到基于甲醇的生物合成的极高的需氧量以及合成气体和氧气的气态底物进入液体培养介质的相对缓慢的相变的限制。为了解决这些技术问题，德累斯顿工业大学自然材料技术研究所建议将合成气和甲醇化学转化为液体和非挥发性C2-化合物乙二醇(EG)和乙醇醛(GA)。利用这些液体化合物作为碳源，解决了相变问题。此外，将这些分子转化为增值产品(通过本文提出的合成代谢途径)的需氧量明显低于直接使用甲醇作为底物时的需氧量。各种细菌能够利用EG和GA作为碳源。然而，天然的EG同化途径包含几个释放二氧化碳的反应，这使得它们特别不适合于乙酰辅酶A依赖的产物合成(乙酸辅酶A在EG上的最高碳产率为50%)。在本研究项目中，提出了一种合成代谢途径，使乙二醇乙酰-辅酶A的生物合成能够节省碳。该途径依赖于EG初始氧化为GA以及GA与3-磷酸甘油醛(GAP)的Aldol加成反应。反应产物阿拉伯糖5-磷酸通过几个反应步骤进一步处理以生成乙酰辅酶A并再生GA受体间隙。合成途径的功能将首先通过使用分离的酶在无细胞反应系统中进行演示。这项工作之后，在工程化的大肠杆菌菌株中实施了该途径。在体内，该途径的功能将通过显示乙酰-辅酶A衍生的甲伐他酸的生物合成来证明。系统水平的生理学分析将指导EG同化和甲羟戊酸生产的优化。本研究为开发微生物产品合成的替代碳源做出了重要贡献。