Engineering synthetic pathways to bio-ethylene production in Cupriavidus necator

背景：乙烯目前是由乙烷的蒸汽裂解产生的，其产生大量的CO2，导致全球变暖。2000年，蒸汽裂解的一次能源使用量为30亿千兆焦耳，排放了约2亿吨二氧化碳。乙烯是最常见的塑料聚乙烯的单体，全球年产量约为8000万吨。找到一种可持续的或碳中和的乙烯生产替代品势在必行。钩虫贪铜菌是一种革兰氏阴性土壤细菌，能够在CO2和H2上生长，从而能够以最小的CO2释放到环境中来生产低碳燃料和化学品。该领域的研究处于绿色革命的前沿，从可持续或碳中性来源生产生物乙烯进一步减少了世界各地对化石燃料的依赖。目的：该项目的目的是将钩虫贪铜菌设计成生产乙烯等碳氢化合物产品的平台。作为概念的证明，我们已经表达了来自P. paseolicola，其足以在异源宿主和青枯雷尔氏菌中产生乙烯。我们已经从基本培养基和二氧化碳中产生了乙烯;我们现在正在通过定向进化和代谢工程来提高产量。作为这个过程的一部分，我们想利用植物的Yang途径设计一种乙烯生产的合成途径。这为在C中实施新途径提供了令人兴奋的机会。necator和链接乙烯生产的增长。乙烯由1-氨基环丙烷-1-羧酸（ACC）有效地生物合成（Zhou等人，2002），其本身衍生自作为杨循环分支的甲硫氨酸（Wang et al.，2002年）。该过程在能量上是有效的，因为它保留了高能甲硫氨酸硫醚键。该途径利用SAM转肽酶、ACC合成酶和ACC氧化酶。1-氨基-环丙烷-1-羧酸（ACC）转化为乙烯释放氰基甲酸，氰基甲酸自发地脱羧以释放氰化物，氰化物主要通过CAS途径解毒（machingura等人，2016）。这一途径的实施将提供一个机制，解毒氰化物在C。钩虫培训内容：该项目将允许在一个独特的多学科环境中进行培训，包括基因组工程，气体发酵，合成生物学，尖端分子生物学和系统生物学建模。该项目将为学生提供大量的可转移技能，在不断增长的生物经济中受到雇主的高度重视。该项目还将提供若干影响力大的出版物。该翻译项目将在诺丁汉的BBSRC/EPSRC合成生物学研究中心（SBRC）内进行，该中心由70多名研究生和博士后研究人员组成（www.clostron.com/people.php），目前的预算为2700万英镑。

Background: Ethylene is currently produced from stream cracking of ethane which produces large quantaties of CO2, contributing to global warming. In 2000, steam cracking had a primary energy use of 3 billion Gigajoules and accounted for approximately 200 millions tons of CO2 emissions. Ethylene is the monomer for the most common plastic, polyethylene, and annual global production is approximately 80 million tons. Finding a sustainable or carbon neutral alternative to ethylene production is imperative. Cupriavidus necator is a gram-negative soil bacterium, capable of growing on CO2 and H2 enabling low carbon fuels and chemicals to be produced with minimal release of CO2 to the environment. Research in this area is at the forefront of the green revolution and the production of bio-ethylene from sustainable or carbon neutral sources further spearheads a diminished reliance on fossils fuels throughout the world.Aim: The aim of this project is to engineer Cupriavidus necator as a platform for the production of hydrocarbon-based products such as ethylene. As proof of concept, we have expressed the efe genes (ethylene forming enzyme) from P. syringae pv. paseolicola, which is sufficient for ethylene production in heterologous hosts and Ralstonia solanacearum. We have generated ethylene from minimal media and from CO2; and we are now in the process of improving production through directed evolution and metabolic engineering. As part of this process we would like to engineer a synthetic pathway for ethylene production utlising the Yang pathway from plants. This provides an exciting opportunity to implement a novel pathway in C. necator and link ethylene production to growth. Ethylene is efficiently biosynthesized from 1-aminocyclopropane-1-carboxylic acid (ACC) (Zhou et al., 2002), which is itself derived from methionine as a branch of the Yang cycle (Wang et al., 2002). This process is energetically efficient as it preserves the high-energy methionine thioether bond. This pathway utilises SAM synthtase, ACC synthase and ACC oxidase. The conversion of 1-amino-cyclopropane-1-carboxylic acid (ACC) to ethylene releases cyanoformic acid, which spontaneously decarboxylates to release cyanide, which is principally detoxified by the CAS pathway (machingura et al., 2016). The implementation of this pathway will provide a mechanism for detoxifiying cyanide in C. necator. Training: The project will allow for training in a unique multidisciplinary environment, incorporating genomic engineering, gas fermentation, synthetic biology, cutting edge molecular biology and systems biology modelling. The project will provide the student with a vast array of transferable skills, highly prized by employers in the growing bioeconomy. The project will also provide several high impact publications. This translational project will be carried out within the BBSRC/EPSRC Synthetic Biology Research Centre (SBRC) at Nottingham which comprises 70+ graduate and postdoctoral researchers (www.clostron.com/people.php) and a current budget of £27M.