Development of reaction systems for ATP regeneration from the inexpensive bulk chemical ethylene glycol

在本项目中，将开发用于技术反应系统中成本效益高的ATP再生的酶系统。该项目计划作为Thomas Walther教授（德累斯顿工业大学生物过程工程教授）和Andreas Liese教授（汉堡工业大学技术生物催化主席）工作组的联合项目，其中AG Walther负责ATP再生酶级联的设计和构建以及单个酶活性的优化。AG Liese将致力于反应工程和反应系统的优化以及向更大规模的转移。ATP再生反应系统具有多种工业应用。例子包括磷酸化精细化学品的合成，以磷酸化为中间步骤的合成和无细胞蛋白表达。ATP依赖性、无细胞产物合成商业化的一个主要障碍是用于ATP再生的底物和辅因子有时价格非常高。为了解决这一技术问题，建议开发反应系统，使低成本和潜在的“绿色”散装化学乙二醇的ATP再生成为可能。设计的酶级联的实现需要增强酶在非天然底物上的活性。这种活性的增加是通过合理的和进化的蛋白质设计相结合来实现的。此外，反应技术分析和优化的各种系统在技术规模上的转移进行了审查。用于ATP再生的酶级联将由分离的酶构建，并在全细胞生物催化剂中实现。在每种情况下，atp再生系统耦合到atp消耗反应的合成，以合成磷酸化或非磷酸化的最终产物，以确定两种设计变体的优点和局限性。所描述的反应级联有潜力发展成为一种新的通用ATP再生系统，也可以在工业规模上使用。

In this project, enzyme systems for the cost-effective regeneration of ATP in technical reaction systems will be developed. The project is planned as a joint project of the working groups of Prof. Thomas Walther (Professorship of Bioprocess Engineering, TU Dresden) and of Prof. Andreas Liese (Chair of Technical Biocatalysis, TU Hamburg), where AG Walther is responsible for the design and construction of the ATP regenerating enzyme cascades as well as for the optimization of individual enzyme activities, and the AG Liese will work on the reaction engineering and optimization of the reaction systems as well as transfer to larger scales.ATP regenerating reaction systems have a variety of industrial applications. Examples include the synthesis of phosphorylated fine chemicals, syntheses with phosphorylation as an intermediate step and cell-free protein expression. A major obstacle to the commercialization of ATP-dependent, cell-free, product syntheses is the sometimes very high price of the substrates and cofactors used for ATP regeneration.To solve this technical problem, the development of reaction systems is proposed, which allow an ATP regeneration of the low-cost and potentially "green" bulk chemical ethylene glycol. The implementation of the designed enzyme cascades requires the enhancement of enzyme activity on a non-natural substrate. This increase in activity is to be achieved through a combination of rational and evolutionary protein design. In addition, a reaction-technical analysis and optimization of the transfer of the various systems on a technical scale is examined.The enzyme cascades for the regeneration of ATP will be constructed from isolated enzymes as well as be implemented in whole-cell biocatalysts. In each case, the ATP-regenerating systems are coupled to the synthesis of an ATP-consuming reaction for the synthesis of a phosphorylated or non-phosphorylated end product in order to determine advantages and limits of both design variants.The reaction cascades described have the potential to be developed into a new universal ATP regeneration system, which can also be used on an industrial scale.