Optimization of a blueprint for in vitro glycolysis by metabolic real-time analysis
Optimization of a blueprint for in vitro glycolysis by metabolic real-time analysis
复制标题
DOI：
10.1038/nchembio.541
复制
发表时间：
2011-05-01
影响因子：
14.8
通讯作者：
Panke, Sven
中科院分区：
文献类型：
--
作者：
Bujara, Matthias;Schuemperli, Michael;Panke, Sven
招募复杂的代谢反应网络用于化学合成已经引起了相当大的关注，但经常需要优化网络组成和动力学以达到足够的生产率。由于预测网络中所有酶的最佳水平的设计框架目前不可用，最先进的途径优化依赖于高通量表型筛选。我们在这里提出了一种新的体外实时分析方法的开发和应用，用于综合研究和合理规划酶网络的合成任务。我们首先利用这一点来合理和快速地推导出一个最佳的蓝图，用于通过大肠杆菌的高度进化的糖酵解来生产精细化学品的结构单元磷酸二羟丙酮（DHAP）。其次，该方法指导了蓝图的三步遗传实施，产生了一个合成操纵子，其预测的DHAP糖酵解通量增加了2.5倍。新的分析设置大大加速了合成多酶网络的合理优化。
Recruiting complex metabolic reaction networks for chemical synthesis has attracted considerable attention but frequently requires optimization of network composition and dynamics to reach sufficient productivity. As a design framework to predict optimal levels for all enzymes in the network is currently not available, state-of-the-art pathway optimization relies on high-throughput phenotype screening. We present here the development and application of a new in vitro real-time analysis method for the comprehensive investigation and rational programming of enzyme networks for synthetic tasks. We used this first to rationally and rapidly derive an optimal blueprint for the production of the fine chemical building block dihydroxyacetone phosphate (DHAP) via Escherichia coli's highly evolved glycolysis. Second, the method guided the three-step genetic implementation of the blueprint, yielding a synthetic operon with the predicted 2.5-fold-increased glycolytic flux toward DHAP. The new analytical setup drastically accelerates rational optimization of synthetic multienzyme networks.