Systems metabolic engineering of Hydrogenophaga pseudoflava for aerobic biosynthesis of fatty acids using CO2 and electron carriers in a novel bioelectrochemical system

电生物技术是电化学和生物技术的一种很有前途的技术，在生物电化学系统（BES）中利用CO2和电力进行化学品和燃料的微生物生物合成。在这方面，一种有吸引力的方法是将CO2转化为CO并电化学形成合成气（H2，CO，CO2）;然后将后者生物转化为化学品或液体燃料。尽管合成气的解耦生物转化最近取得了令人印象深刻的进展，但该技术仍然存在一些固有的障碍，例如气体到培养基中的有限传质，电子或电子载体到微生物宿主的低吸收或低效转移，以及关于底物的毒性和爆炸性的安全问题。此外，产乙酸菌作为用于合成气生物转化的主要使用的微生物宿主具有有限的产物谱，因为更复杂分子的产生在代谢能力之外。在本项目中，拟黄氢噬菌体在合成气好氧利用方面的巨大潜力和工程化H。pseudoflava用于生产脂肪酸将在一种新的直接电微生物生产系统中进行探索，该系统具有原位和按需生产H2和O2（来自水）和CO（来自CO2）。开发一个系统的和定量的了解电子转移及其对氧化还原和能量代谢的一氧化碳营养细菌的影响，我们将应用代谢组学，通量分析和量化动力学参数的野生型和工程突变体缺陷的电子转移，能量和氧化还原代谢下给定的气体组合物提供了一个优化的BES。BES将解决生物电化学系统和气体发酵目前的局限性，如上所述。本论文将利用所获得的知识建立第一个H. pseudoflava，特别是关于吸收不同的电子载体，能量和氧化还原平衡。此外，我们将工程师H。pseudoflava用于生产代表ATP和NADPH密集型产品类别的脂肪酸，因此其过量生产将挑战H.特别是在自养条件下。开发菌株的定量分析将用于评估和完善代谢和电子转移模型。该项目将开辟新的可能性，工程有效的电微生物生产菌株，并开发改进的电发酵。

Electrobiotechnology is a promising technology at the interface of electrochemistry and biotechnology to use CO2 and electricity for the microbial biosynthesis of chemicals and fuels in bioelectrochemical systems (BES). An appealing approach in this regard is the conversion of CO2 to CO and the formation of syngas (H2, CO, CO2) electrochemically; the latter is then biologically converted into chemicals or liquid fuels. Although the decoupled bioconversion of syngas has made impressive progresses recently, this technology still has several inherent obstacles such as limited mass transfer of gases into culture medium, low uptake or inefficient transfer of electrons or electron carriers to the microbial host, and safety issues regarding toxicity and explosiveness of the substrates. Furthermore, acetogens as the mostly used microbial hosts for syngas bioconversion have a limited spectrum of products since the production of more complex molecules is outside the metabolic capacity. In this project, the great potential of Hydrogenophaga pseudoflava in the aerobic utilization of syngas and the capacity of engineered H. pseudoflava for the production of fatty acids will be explored in a novel direct electromicrobial production system with in situ and on demand production of H2 and O2 (from water) and CO (from CO2). To develop a systemic and quantitative understanding of the electron transfer and its impact on redox and energy metabolism of this carboxydotrophic bacterium, we will apply metabolomics, flux analysis and quantify kinetic parameters of the wild type and engineered mutants defective in electron transfer, energy and redox metabolism under given gas compositions provided by an optimized BES. The BES will address current limitations of bio-electrochemical systems and gas fermentations as already mentioned above. The gained knowledge will be utilized to set up a first metabolic and electron transfer model of the autotrophic metabolism of H. pseudoflava, especially regarding uptake of the different electron carriers, energy and redox balances. Furthermore, we will engineer H. pseudoflava for the production of fatty acids which represent an ATP and NADPH intensive product class and therefore its overproduction will challenge the metabolism of H. pseudoflava especially under autotrophic conditions. A quantitative analysis of the developed strains will be used to evaluate and refine the metabolic and electron transfer model. This project will open up new possibilities to engineer efficient electromicrobial production strains and to develop improved electro-fermentation.