大肠杆菌超氧化应激时赖氨酸乙酰化修饰网络的化学热动力学状态对中心碳代谢系统的调控作用

本项目旨在探究超氧化应激情况下赖氨酸乙酰化修饰在代谢流量重分配过程中的调控作用，同时部分解释单一（去）乙酰化体系对众多不同底物的差异化调控机制。本项目假说认为超氧化应激会对（去）乙酰化体系的化学热动力学状态产生扰动,驱动代谢酶乙酰化修饰谱改变，最终对代谢流量重分配产生贡献。为此，我们利用代谢物组学手段,mRNA检查和酶学手段，解析超氧化应激对（去）乙酰化体系的化学动力学（酶活）和热力学扰动，测量（去）乙酰化体系的动力学和热力学参数，特别是（去）乙酰化酶的Km，Kcat和Kd；利用亲和标签融合和质谱技术分析乙酰化修饰谱改变的原因；利用13C代谢流量验证乙酰化修饰在代谢流量重分配过程中的调控作用。本项目有助于了解乙酰化对于中心代谢调控的机理,对于改造和设计具有特定流量的代谢网络有重要意义;本项目从新视角认识氧化应激的致病机理，为氧化应激相关疾病的诊疗提供新的思路，靶点和途径。

The purpose of this project is to explore the regulatory role of lysine acetylation modification in metabolic flux redistribution process under paraquat induced oxidative stress. In addition,the projct aims to reveal the underlying mechanism of different acetylation on the various substrates by the sole acetylation systems.The project holds a conjecture that oxidative stress would exert disturbance on both chemical dynamic and thermodynamic state of acetylation /deacetylatton system, thereby driving the coordinated changes in acetylation states of the various metabolic enzymes.The conjecture also assumed the changes in acetylation state contribute to metabolic flux redistribution in response to the oxidative stress. The pertubation of acetylation system as well as its dynamic and thermodynamic constants (Km,Kcat and possible Kd)were explored by means of metabolomics analysis, mRNA detection method and enzymatic method.Then,we utilize epitope tags fusion techniques, mass spectrometry and proteomic method to analyze the resulting effect on the acetylation spectra. Afterwards, we will use 13C metabolic flux to validate the actual role of acetylation in metabolic flux redistribution process. This project aims at understanding the regulatory mechanism of acetylation on metabolic flux and would be meaningful in renovation and design of particular metabolic network. Besides, this project help to understand the disruption of oxidative stress on the the acetylation process, providing a new perspective about the pathogenic mechanism of oxidative stress,thus would contribute to screening new targets and finding new treatments for oxidative stress-related diseases. This project has been of interest and significance to both the systems biology and medical communities.