A combinatorial assembly strategy to optimise biosynthetic pathway performance in yeast - a synthetic biology

微生物生物合成途径是高价值产品的重要来源，包括工业前体、农用化学品、药物、食品添加剂、生物燃料和生物制药，也可以就地应用于生物修复、生物过滤、生物质生产和其他过程。然而，为了优化生产力，通常需要在“路径开发”上投入大量的时间和精力。由于任何给定的生物合成途径的生产力都受到遗传因素、宿主生理、代谢动力学和速率控制特性的影响，因此通过传统手段对生物合成途径进行优化可能是一个偶然的过程，浪费了时间和资源。在这里，我们专注于酿酒酵母，这是一种GRAS（通常被认为是安全的）生物，具有成熟的生物技术血统，但我们的策略同样可以应用于其他微生物。由于合成生物学和系统生物学领域的实验和理论方法的快速发展，以及DNA合成成本的迅速下降，我们将在这个项目中遵循的策略直到最近才变得可行。例如，我们现在可以利用计算设计算法、机器人技术、DNA合成和快速多组分DNA组装技术来并行探索大量的生物合成途径变异。既然这些新的机会对我们来说是可用的，我们就可以利用它们来绕过传统的途径开发的试错方法。通过构建和优化模型（纤维素降解酶）途径，证明具有成本效益的目标分子（EtOH）的生产将代表Ingenza在生物燃料领域的商业机会。此外，更有效地将原料转化为有价值的化学品可能带来中长期的环境效益。

Microbial biosynthetic pathways are an important source of high value products, including industrial precursors, agrochemicals, drugs, food additives, biofuels and biopharmaceuticals, and can also be applied in situ for bioremediation, biofiltration, biomass production and other processes. However, a considerable amount of time and effort generally have to be invested in 'pathway development' in order to optimise productivity. Since the productivity of any given biosynthetic pathway is influenced by genetic factors, host physiology, metabolic dynamics and rate control characteristics, the optimization of biosynthetic pathways by traditional means can be a hit-and-miss process that wastes time and resources. Here, we focus on Saccharomyces cerevisiae, which is a GRAS (generally regarded as safe) organism with a proven biotechnological pedigree, but our strategy could equally be applied to other microbes. The strategy that we will follow in this project has only very recently become feasible by virtue of rapid progress in experimental and theoretical methods in the fields of synthetic and systems biology and also due to the rapidly decreasing cost of DNA synthesis. For example, we can now explore very large numbers of biosynthetic pathway variants in parallel utilizing computational design algorithms, robotics, DNA synthesis and rapid multi-component DNA assembly technologies. Now that these new opportunities are available to us, we can use them to bypass the traditional trial-and-error approach to pathway development. Demonstration of cost-effective production of the target molecule (EtOH) through construction and optimisation of the model (cellulose-degrading enzyme) pathway will represent a commercial opportunity in the biofuel field for Ingenza. In addition, more efficient feedstock conversion into valuable chemical(s) could result in mid- to long-term environmental benefits.