小分子介导的位点选择性PGAM1赖氨酸乙酰化研究

Protein post-translational modification is an important signal transduction in living system. The ability of chemists to install these covalent additions selectively has been critical for elucidating their roles in biology. Phosphoglycerate mutase 1(PGAM1) is a glycolitic enzyme which converts 3PG to 2PG. Its post-translational modification plays an important role in biological function. To study the biological function of acetylation in PGAM1, we proposed a strategy that a small molecule PGAM1 inhibitor being as war-head, transferred its reactive acetyl group to the specific lysine near the binding site to achieve site selective modification. Based on our previous studies of SAR and co-crystal structure, we designed and sythesized the lead compound Ac-1, which can effectively acetylate the lysine (K100) of PGAM1 in vitro and in vivo. In this study, we will further optimize the chemical structure of lead compound based by introducing deuterium and click group into reactive group for further quantitation and target validation. The acetylation will be validated by chemical biology methods including high-resolution mass spectrometry, crystallography to elaborate the mechanism and function. Our aim is to develop a class of biorthogonal and site-specific PGAM1 acetylation small molecule, which could finally clarify the biological function and mechanism of PGAM1 post-translational modification.

蛋白质翻译后修饰是生物体内一种重要的信号传导方式，通过化学手段选择性修饰特定氨基酸对阐述其生物学功能极其重要。磷酸甘油酸变位酶1(PGAM1)是细胞糖酵解途径中的酶，它的乙酰化修饰具有重要的生物学功能。为研究其乙酰化功能，本课题提出一种小分子介导的蛋白位点选择性乙酰化策略，即以PGAM1小分子抑制剂为靶头，携带可反应的乙酰基团靶向转移给结合位点附近的特定赖氨酸后并离去，实现位点选择性乙酰化修饰。申请人通过结构导向性策略设计化学探针Ac-1，在体外和活细胞水平能够选择性乙酰化目标赖氨酸K100。本课题将对先导探针进行结构修饰，分别引入氘代和click基团来研究体内乙酰化效率和脱靶效应，后续通过多种化学生物学手段表征该类修饰的细胞内有效性和靶向性，结构生物学手段解释共价修饰机制。本课题将聚焦于采用化学手段对活细胞单一蛋白PGAM1进行实时动态的乙酰化调控，阐明PGAM1翻译后修饰的生物学意义。

本项目聚焦于采用化学手段对活细胞单一蛋白PGAM1进行实时动态的乙酰化调控，阐明PGAM1翻译后修饰的生物学意义。本研究在充分探索PGAM1抑制剂构效关系的基础上，完成了乙酰化小分子KHAc的设计合成，并在分子水平进行了小分子介导的蛋白质赖氨酸乙酰化的确认，利用结构生物学手段揭示了乙酰化反应的动态过程；随后在活细胞上表征了小分子靶向性调控PGAM1的100位赖氨酸乙酰化，揭示了其乙酰化后的能够影响细胞F-actin生成以及细胞迁徙能力的生物学功能。本项目的研究成果已发表基金标注的 SCI 收录文章5篇，包括 Cell Metabolism 及PNAS，申请中国发明专利一项，PGAM1的抑制剂成功转让至企业进行进一步合作开发，获得中国药学会药学科技二等奖和上海市药学会药学科技一等奖。