Quiescent Microbial Cell Factories

人们普遍认识到，化学品和燃料的生产必须从石化产品转向可持续的原料基地。然而，越来越明显的是，燃料和化学品的生物制造工艺的开发将需要能够提高工艺生产率、稳健性和经济竞争力的技术改进。改进的生物制造的一个关键组成部分是改进的细菌细胞工厂的开发，这构成了这一应用的焦点。细菌细胞工厂开发的一个普遍接受的标准是，它必须将最大量的原材料直接转化为所需的最终产品，副产品积累最少。虽然许多研发工作致力于消除不需要的代谢产物的共同生产，但由于微生物生长而积累的主要副产品细胞生物量通常被视为该过程的不可避免的后果。然而，通过将产物形成与生长分离，保持生物量恒定，同时允许底物转化为产物持续延长的时间段，可以提高效率。这种方法已被利用在生产某些氨基酸的转基因菌株，但并不普遍适用于大多数的代谢工程策略，其中生长是必需的，以保持代谢健康的细菌cell.The重点的这个项目是使用静止细胞（Q-Cell）技术开发的萨默斯实验室在遗传学系在剑桥大学。Q-Cells是由大肠杆菌产生的非生长但代谢活跃的细胞工厂。具有特定遗传修饰的细菌宿主可以通过添加吲哚作为静止的化学触发物而被诱导进入这种状态。在实验室研究中，Q-细胞的生产率比传统的E-细胞高10倍。大肠杆菌培养物，在没有生物质产生的情况下，资源被更有效地引导到产品中。此外，中心碳代谢的途径通量在静止期开始后仍然很高，提供了一个不断再生的代谢产物池，可以转化为product.The性能的Q-电池在相关的工业应用条件下将在这个项目中进行评估。在剑桥大学遗传学系，将在实验室规模上改进该系统，比较一系列化学触发剂诱导静止的功效。还将评估环境因素（温度）对系统有效运行的作用。希望这些变化可以避免对细菌宿主菌株进行遗传修饰的需要，从而增加系统的易用性并提高其效率。在剑桥的研究将在摇瓶培养中进行，也可能在小规模发酵罐中进行。然而，必须确定在这些条件下的良好性能是否会扩大到工业条件。这就是工业伙伴CPI的作用至关重要的地方。CPI最初将对目前形式的Q-Cell系统的效用进行严格测试。由于在剑桥的工作表明了潜在的改进，这些改进将被纳入CPI计划。

It is widely recognised that the manufacture of commodity chemicals and fuels must switch from petrochemicals to a sustainable feedstock base. However it is also increasingly evident that development of processes for the bio-manufacture of fuels and chemicals will require technology improvements capable of increasing process productivity, robustness and economic competitiveness. A key component of improved bio-manufacture is the development of improved bacterial cell factories, which constitutes the focus of this application.A universally accepted criterion for a bacterial cell factory development is that it must convert the maximum amount of raw material directly to the desired end product with minimum by-product accumulation. While much R&D effort is dedicated to eliminating the co-production of undesirable metabolites, the primary by-product, cell biomass accumulated due to microbial growth, is often dismissed as an inevitable consequence of the process. However, efficiency would be increased by uncoupling product formation from growth, keeping biomass constant while allowing the conversion of substrate to product for an extended period of time. This approach has been exploited in the production of certain amino acids by genetically modified strains but is not generally applicable for the majority of metabolic engineering strategies where growth is required to maintain the metabolic health of the bacterial cell.The focus of this project is the use of quiescent cell (Q-Cell) technology developed in the Summers laboratory in the Department of Genetics at Cambridge University. Q-Cells are a non-growing but metabolically-active cell factory generated from the bacterium Escherichia coli. The bacterial host, possessing a specific genetic modification, can be induced into this state by the addition of indole as a chemical trigger of quiescence. In laboratory studies the productivity of Q-cells is up to 10-fold greater than conventional E. coli cultures, resources being channelled more efficiently into product in the absence of biomass generation. Moreover, pathway flux in central carbon metabolism remains high after the onset of quiescence, providing a constantly regenerated pool of metabolites that can be diverted into product.The performance of Q-cells under conditions relevant to industrial application will be evaluated in this project. At Cambridge University Department of Genetics, work will be undertaken to improve the system on the laboratory scale, comparing the efficacy of a range of chemical triggers to induce quiescence. There will also be an assessment of the role of environmental factors (temperature) on the efficient operation of the system. It is hoped that these changes might avoid the need for genetic modification of the bacterial host strain, thus increasing the ease of use of the system as well as increasing its efficiency. The investigations at Cambridge will be conducted in shake-flask culture and possibly in small-scale fermenters. However it is essential to determine whether good performance under these conditions will scale up to industrial conditions. This is where the role of the industrial partner CPI is crucial. CPI will initially conduct rigorous testing of the utility of the Q-Cell system in its present form. As work in Cambridge suggests potential improvements these will be incorporated into the CPI programme.