Department of Bioengineering and Therapeutic sciences, University of California, San Francisco, San Francisco, CA, USA.
Department of Chemistry, University of Michigan, Ann Arbor, MI, USA.
Nat Commun. 2021 Nov 23;12(1):6803. doi: 10.1038/s41467-021-27185-9.
Enzymes are represented across a vast space of protein sequences and structural forms and have activities that far exceed the best chemical catalysts; however, engineering them to have novel or enhanced activity is limited by technologies for sensing product formation. Here, we describe a general and scalable approach for characterizing enzyme activity that uses the metabolism of the host cell as a biosensor by which to infer product formation. Since different products consume different molecules in their synthesis, they perturb host metabolism in unique ways that can be measured by mass spectrometry. This provides a general way by which to sense product formation, to discover unexpected products and map the effects of mutagenesis.
酶在广阔的蛋白质序列和结构形式空间中都有体现，其活性远远超过了最好的化学催化剂；然而，由于缺乏用于检测产物形成的技术，对它们进行具有新颖或增强活性的工程改造受到了限制。在这里，我们描述了一种通用的、可扩展的酶活性表征方法，该方法利用宿主细胞的代谢作为生物传感器，通过该传感器来推断产物的形成。由于不同的产物在合成过程中消耗不同的分子，它们会以独特的方式干扰宿主代谢，这些可以通过质谱来测量。这为检测产物形成、发现意想不到的产物以及绘制诱变影响提供了一种通用的方法。
