Department of Bioengineering, University of California, San Diego, La Jolla, CA 92093.
The Novo Nordisk Foundation Center for Biosustainability, University of California, San Diego, La Jolla, CA 92093.
Proc Natl Acad Sci U S A. 2018 Oct 23;115(43):11096-11101. doi: 10.1073/pnas.1811971115. Epub 2018 Oct 9.
Understanding the complex interactions of protein posttranslational modifications (PTMs) represents a major challenge in metabolic engineering, synthetic biology, and the biomedical sciences. Here, we present a workflow that integrates multiplex automated genome editing (MAGE), genome-scale metabolic modeling, and atomistic molecular dynamics to study the effects of PTMs on metabolic enzymes and microbial fitness. This workflow incorporates complementary approaches across scientific disciplines; provides molecular insight into how PTMs influence cellular fitness during nutrient shifts; and demonstrates how mechanistic details of PTMs can be explored at different biological scales. As a proof of concept, we present a global analysis of PTMs on enzymes in the metabolic network of Based on our workflow results, we conduct a more detailed, mechanistic analysis of the PTMs in three proteins: enolase, serine hydroxymethyltransferase, and transaldolase. Application of this workflow identified the roles of specific PTMs in observed experimental phenomena and demonstrated how individual PTMs regulate enzymes, pathways, and, ultimately, cell phenotypes.
理解蛋白质翻译后修饰（PTMs）的复杂相互作用是代谢工程、合成生物学和生物医学科学的主要挑战。在这里，我们提出了一个工作流程，该流程集成了多重自动化基因组编辑（MAGE）、基因组规模代谢建模和原子分子动力学，以研究 PTM 对代谢酶和微生物适应性的影响。该工作流程整合了跨学科的互补方法；提供了关于 PTM 如何在营养物质转移过程中影响细胞适应性的分子见解；并展示了如何在不同的生物学尺度上探索 PTM 的机制细节。作为概念验证，我们对代谢网络中酶的 PTM 进行了全局分析。基于我们的工作流程结果，我们对三种蛋白质中的 PTM 进行了更详细的、机制上的分析：烯醇酶、丝氨酸羟甲基转移酶和转醛醇酶。该工作流程的应用确定了特定 PTM 在观察到的实验现象中的作用，并展示了单个 PTM 如何调节酶、途径，最终调节细胞表型。
