乙醛脱氢酶2介导的4-羟基壬烯醛蛋白修饰位点鉴定及其生物学效应研究

4-hydroxynonenal is the most abundant and toxic end-products of Lipid peroxidation caused by oxidation damage, the protein adducts of which may cause imflamation, cell death and so on. Recently, our study suggested that aldehyde dehydrogenase (ALDH2) plays pivtol role in protecting renal cells from oxidation damage caused by chemical therapy compound doxorubicin by catalyzing 4-HNE into non toxic 4-hydroxynonanoic acid. Identification of 4-HNE protein modified site provides confidential evidence to explain the function of the modification on the protein and its biological effects. However, the enrichment method still need to be improved, while the mechnsim and the molecular targets of 4-HNE has not been clarfied sysmatically. For this purpose, the applicant has developed nanoparticles to capture 4-HNE modified peptides from complex samples. On these basis, the present project will try to enrich and identify 4-HNE modified peptides under oxidation damage, then constructs oxidation stress signal transduction networks with ALDH2 as the core regulator. Further, the project will discover the relationship between biological effects of 4-HNE modification and ALDH2. The present project will devote the effect to clarify the molecular mechanism of ALDH2 in protecting the cells from oxidation damage. It has the potential to find treatment targets and clinical biomarkers. It is of great significance to promote the knowledge of disease pathogenesis and basic research on therapy.

4-羟基壬稀醛 (4-HNE)是细胞脂质过氧化过程中含量丰富、毒性最强的代谢末产物，其蛋白加成物可引起炎症、细胞死亡等损伤性病变。最近，本课题组发现，乙醛脱氢酶2 (ALDH2) 通过对4-HNE氧化代谢，在阿霉素引起的肾细胞氧化损伤过程中起关键性保护作用。确定4-HNE蛋白质修饰化位点将为研究ALDH2通过下调4-HNE发挥氧化损伤调节作用的分子机制提供重要线索。针对4-HNE修饰化富集方法尚不健全、靶蛋白和分子机制未系统阐明的现状，申请人发展了基于磁性纳米粒子的肽段富集策略。在此基础上，本项目拟通过生物质谱鉴定4-HNE蛋白质修饰位点，构建以ALHD2为调控核心的4-HNE信号转导通路，结合生物学实验探讨4-HNE修饰的生物学效应与ALDH2之间的关系。本项目有望阐明ALDH2对氧化损伤保护作用的分子机制，为疾病治疗提供潜在药物靶点及临床诊断标志物，对推动疾病发病学研究具有重要意义。

4-羟基壬稀醛 (4-HNE)是细胞脂质过氧化过程中含量丰富、毒性最强的代谢末产物，其蛋白加成物可引起炎症、细胞死亡等损伤性病变。最近，本课题组发现，乙醛脱氢酶2 (ALDH2) 通过对4-HNE 氧化代谢，在阿霉素引起的肾细胞氧化损伤过程中起关键性保护作用。确定4-HNE 蛋白质修饰化位点将为研究ALDH2 通过下调4-HNE 发挥氧化损伤调节作用的分子机制提供重要线索。.针对4-HNE 修饰化富集方法尚不健全、靶蛋白和分子机制未系统阐明的现状，申请人发展并优化了基于磁性纳米粒子的肽段富集策略，鉴定了鉴定了氧化损伤条件下细胞中4-HNE修饰蛋白及位点。在此基础上，开展了基于全氟化合物的4-HNE修饰位点富集方法学探索。探索了市售全氟化物对4-HNE修饰肽段的反应、富集条件研究，并合成全氟化肼探针进行衍生化反应探索。.ALDH2对氧化应激的调控作用与4-HNE修饰具有相关性。通过动物实验，明确了乙醛脱氢酶2对氧化损伤的调节作用受VHL蛋白调控。进一步地，通过分子机制研究，初步探讨了VHL通过非HIF-1a依赖的信号通路及HNF-4a转录调控对ALDH2介导的氧化损伤的进行调控。此项研究为疾病治疗提供潜在药物靶点及临床诊断标志物，对推动疾病发病学研究具有重要意义。 .