蛋白质泛素化靶向识别与动态修饰分析

Ubiquitination, as a dynamic multifaceted post-translational modification, plays an important role in regulating protein structures and functions. Imbalance and abnormality in the dynamic process of ubiquitination are closely related to various diseases. Given the molecular features of ubiquitination in diversity and multi-valence, herein design and screen of targeting peptides with high affinity and specificity towards ubiquitin dimers is proposed. Ubiquitin dimers with characteristic conformations are chosen as the targets. Materials for affinity separation will be developed by using peptides as recognition ligands. Based on the specific interactions between peptides and ubiquitin dimers, novel methods for separation and enrichment of ubiquitinated proteins will be established. Bioprobes with peptide recognition-induced emission will be designed and synthesized. Analytical methods for imaging, detecting and tracing protein ubiquitination in living cells will be developed. The peptide-based materials and bioprobes will be applied to monitor the ubiquitination processes in lesion cells or upon drug treatment. By providing both in situ analysis and off-line sample preparation methods, this study is promising for understanding protein functions, discovering new drug targets and regulating diseases at molecular level.

泛素化修饰是蛋白质结构与功能的重要调控方式，具有高度动态和复杂多变的特点，其动态失衡和异常则与众多疾病的发生发展密切相关。针对泛素化修饰多样性和多价性的分子特征，本项目拟以具有特征构型的泛素二聚体为识别标签，开展高亲和力、高选择性靶向多肽的设计筛选；构建以多肽为界面识别分子的亲和分离材料，建立基于多肽-泛素标签特异性相互作用的泛素化蛋白质分离富集新方法；构建多肽识别荧光响应的生物探针，建立活细胞内蛋白质泛素化的成像检测、示踪分析新方法。利用分离材料和多肽探针，探测病变和药物刺激过程中，蛋白质泛素化动态修饰特征，为蛋白质功能的认识、药物新靶标的发现和化学干预提供新工具和新方法。

泛素化修饰位点复杂、构型多样和高度动态的特点是蛋白质泛素化准确识别检测和调控的关键难题。本项目以具有特征构型的泛素二聚体为识别标签，从天然泛素结合域出发，设计合成了高亲和力、高选择性的小分子多肽；开展了连接骨架分子的设计筛选，获得了空间长度适配的螺旋肽基分子，组装得到与靶标泛素二聚体高特异性的协同型多肽；发现并解析了可见光响应的多肽探针顺-反异构现象，为动态多变的蛋白质泛素化修饰的识别和原位操控提供特异性新工具。以多肽为界面识别分子，制备了核壳型亲和分离纳米材料和多肽功能化仿生亲和界面，基于可逆共价键和磁操控，实现了复杂生物体系中蛋白质靶向分离富集。构建了特征泛素链识别荧光响应生物探针，建立了蛋白质泛素化修饰原位成像检测和示踪分析新方法；在活细胞和活体水平，实现了自噬和热应激过程蛋白质泛素化修饰含量和时空变化差异分析。利用线粒体自噬过程干预探针，实现了恶性肿瘤的高效抑制；为蛋白质功能的认识、药物靶标的发现和化学干预提供新工具和新方法。本项目在Angew. Chem. Int. Ed., Anal. Chem.等学术期刊发表论文9篇，申请中国发明专利2项。项目执行期间，项目负责人2021年获国家自然科学基金优秀青年基金项目资助，获中科院化学所青年科学奖特别优秀奖2项，获中科院化学所青年科学奖优秀奖1项。