Jeschek Markus, Gerngross Daniel, Panke Sven
Department of Biosystems Science and Engineering, ETH Zurich, Basel 4058, Switzerland.
Nat Commun. 2016 Mar 31;7:11163. doi: 10.1038/ncomms11163.
Rational flux design in metabolic engineering approaches remains difficult since important pathway information is frequently not available. Therefore empirical methods are applied that randomly change absolute and relative pathway enzyme levels and subsequently screen for variants with improved performance. However, screening is often limited on the analytical side, generating a strong incentive to construct small but smart libraries. Here we introduce RedLibs (Reduced Libraries), an algorithm that allows for the rational design of smart combinatorial libraries for pathway optimization thereby minimizing the use of experimental resources. We demonstrate the utility of RedLibs for the design of ribosome-binding site libraries by in silico and in vivo screening with fluorescent proteins and perform a simple two-step optimization of the product selectivity in the branched multistep pathway for violacein biosynthesis, indicating a general applicability for the algorithm and the proposed heuristics. We expect that RedLibs will substantially simplify the refactoring of synthetic metabolic pathways.
在代谢工程方法中，合理的通量设计仍然困难，因为重要的途径信息常常无法获得。因此，采用了经验方法，即随机改变途径中酶的绝对和相对水平，随后筛选性能得到改善的变体。然而，筛选在分析方面往往受到限制，这强烈促使人们构建小型但精巧的文库。在此，我们引入RedLibs（精简文库），这是一种算法，可用于合理设计用于途径优化的智能组合文库，从而最大限度地减少实验资源的使用。我们通过荧光蛋白的计算机模拟和体内筛选，证明了RedLibs在核糖体结合位点文库设计中的实用性，并对紫罗碱生物合成的分支多步途径中的产物选择性进行了简单的两步优化，表明该算法和所提出的启发式方法具有普遍适用性。我们期望RedLibs将大大简化合成代谢途径的重构。
