Collaborative Research: A Systems Biology Approach for Metabolically Engineering Escherichia coli for Producing Hydrogen via Fermentation

无害细菌可用于生产供人类使用的化学品，包括燃料。氢分子是一种环保的燃料；因此，如果氢气可以通过细菌有效地合成，那么我们就可以减少空气污染，减少我们对外国石油的依赖，并避免温室气体（我们的过程中形成的二氧化碳将被回收到植物中，并再次用于形成氢气，因此没有净排放）。这项研究将使氢作为一种清洁的能量形式从细菌中产生，利用单糖发酵。在氢气的发酵生产中，葡萄糖转化为丙酮酸，丙酮酸转化为甲酸盐，甲酸盐再转化为氢。因此，必须通过阻止竞争性葡萄糖反应和将丙酮酸转化为醋酸盐、乳酸盐和乙醇或去除甲酸盐的反应，将细菌细胞中的碳通量导向氢的形成。为了实现这一目标，将使用系统生物学方法来产生更多的氢，包括(i)通过去除不需要的途径来改变细菌的代谢，（ii）使用蛋白质工程进化必要的酶，以及（iii）模拟细菌细胞的所有代谢途径，以便产生更好的细菌。我们选择大肠杆菌作为我们的模型系统，因为(i)大肠杆菌BW25113菌株的所有3985个非致死单突变都是可用的；（ii）我们可以使用新的、连续的、基于病毒的方法快速组合突变；（iii）大肠杆菌是研究得最好的细菌，因此对其代谢有很好的了解；（iv）对其基因组进行测序，因此可以使用DNA微阵列，并且可以在基因组尺度上进行代谢通量分析。除了研究成果外，参与这项研究的三位教授还将培训本科生和研究生，通过万维网将该项目的具体成果公之于众，并通过播客将部分研究成果传播给不同的受众。

CBET-0753702WoodNon-harmful bacteria may be used to produce chemicals for humans and this includes fuels. Molecular hydrogen is an environmentally-clean fuel; hence, if hydrogen can be synthesized efficiently by bacteria, then we may reduce air pollution, reduce our dependence on foreign oil, and avoid greenhouse gases (the carbon dioxide formed by our process will be recycled to plant mass and used again to form hydrogen so there are no net emissions). This research will enable hydrogen to be produced as a clean form of energy from bacteria using fermentation of simple sugars. In fermentative production of hydrogen, glucose is converted into pyruvate which is converted to formate which is then converted to hydrogen. Hence, it is imperative to direct the carbon flux in the bacterial cell to hydrogen formation by preventing competing glucose reactions and those that convert pyruvate to acetate, lactate, and ethanol or that remove formate. To achieve this aim, a systems biology approach will be used to produce more hydrogen that includes (i) altering the metabolism of the bacteria by removing unwanted pathways, (ii) evolving the necessary enzymes using protein engineering, and (iii) modeling all the metabolic pathways of the bacterial cell so that even better bacteria may be generated. We have chosen Escherichia coli as our model system since (i) all 3985 non-lethal single mutations of strain E. coli BW25113 are available; (ii) we may combine mutations rapidly using our novel, successive, virus-based method; (iii) E. coli is the best-studied bacterium so its metabolism is well understood; and (iv) its genome is sequenced so DNA microarrays are available and metabolic flux analysis may be carried out at the genome-scale. Along with the research results, the three professors involved in this research will train undergraduate and graduate students, will make accessible, via the World-Wide Web, specific outcomes of the project, and will disseminate some of the research results to a diverse audience using podcasting.