Metabolic adaptability of E. coli: engineering autotrophic CO2 fixation

描述（由申请人提供）：关于中心碳代谢的丰富知识使得生物体能够被改造为生产有用的分子，并提供了对许多疾病的深入了解，包括肥胖、糖尿病、动脉粥样硬化和一些癌症 (1, 2)。代谢工程的进一步进步有可能对环境保护和人类健康产生重大影响。例如，随着人类碳基燃料消耗的增加，大气中的二氧化碳含量不断上升 (3, 4)，预计 50 年的时间尺度上的后果将是巨大的。该提案的目标是通过确定生物体从异养生长模式转变为自养生长模式所需的变化来提高我们对中心碳代谢的理解。具体来说，我建议使用卡尔文循环将已充分表征的异养细菌大肠杆菌转变为自养生长模式，其中将二氧化碳固定为其唯一碳源。这将涉及外源基因的引入、内源基因的靶向突变、代谢通量的定量建模和测量以及定向进化。研究结果将提供有关中心碳代谢的调节、演化和可塑性的信息，并可能为从大气中固碳提供新的方法。 公共卫生相关性：进一步了解大肠杆菌代谢网络如何适应重大变化将至少带来双重好处。首先，我们希望阐明细菌代谢如何适应以补偿碳利用关键步骤的重大变化。其次，我们希望利用这些知识来推动合理改造的大肠杆菌菌株的开发，该菌株能够仅靠二氧化碳生存，从而将大气二氧化碳修复与具有药用或其他有用特性的分子的生产结合起来。

DESCRIPTION (provided by applicant): The wealth of knowledge about central carbon metabolism has allowed organisms to be engineered for the production of useful molecules and has provided insight into many diseases, including obesity, diabetes, atherosclerosis, and some cancers (1, 2). Further advances in metabolic engineering have the potential to significantly impact both environmental protection and human health. For example, CO2 levels in the atmosphere are rising as human consumption of carbon-based fuels increases (3, 4), and the consequences on a 50-year timescale are expected to be dramatic. The goal of this proposal is to improve our understanding of central carbon metabolism by determining the changes needed to convert an organism from a heterotrophic to autotrophic mode of growth. Specifically, I propose to convert the well-characterized heterotrophic bacterium E. coli to an autotrophic mode of growth in which it fixes CO2 as its sole carbon source, using the Calvin cycle. This will involve introduction of foreign genes, targeted mutation of endogenous genes, quantitative modeling and measurement of metabolic fluxes, and directed evolution. The results will provide information about the regulation, evolution and plasticity of central carbon metabolism, and may enable new methods for carbon sequestration from the atmosphere. PUBLIC HEALTH RELEVANCE: Furthering our understanding of how the E. coli metabolic network can adapt to major changes will have at least a two-fold benefit. First, we hope to shed light on how bacterial metabolism can adapt to compensate for major changes to key steps of carbon utilization. Secondly, we hope to use this knowledge to drive the development of a rationally engineered strain of E. coli capable of surviving solely on carbon dioxide for the purpose of coupling atmospheric CO2 remediation with the production of molecules with medicinal or other useful properties.