深圳湾实验室李刚团队报道了使用小分子光敏剂和化学探针中继标记的活细胞功能性组氨酸的全局分析。相关研究成果发表在2024年6月4日出版的《自然—化学》。

化学蛋白质组学的最新进展集中在开发与亲核氨基酸残基反应的化学探针上。尽管组氨酸由于其在酶催化、金属结合和蛋白质-蛋白质相互作用中的重要性而成为一种有吸引力的候选者，但其适度的亲核性带来了挑战。其修饰经常受到半胱氨酸和赖氨酸的影响，导致选择性差和蛋白质组覆盖范围窄。

该文中，研究人员报道了一种对组氨酸具有高选择性的单线态氧和化学探针中继标记方法。筛选了小分子光敏剂和化学探针库，以优化组氨酸标记，使组氨酸能够在具有约7200个独特位点的活细胞中进行分析。利用核磁共振波谱和X射线晶体学，研究人员对反应机理和产物的结构进行了表征。然后，应用这种方法在选定的金属蛋白中发现了，对酶活性和金属结合至关重要的未标记组氨酸位点。

该方法还揭示了由蛋白质-蛋白质相互作用介导的组氨酸的可及性变化，该变化影响选择蛋白质的亚细胞定位，强调了其发现功能组氨酸的能力。

附：英文原文

Title: Global profiling of functional histidines in live cells using small-molecule photosensitizer and chemical probe relay labelling

Author: Zhai, Yansheng, Zhang, Xinyu, Chen, Zijing, Yan, Dingyuan, Zhu, Lin, Zhang, Zhe, Wang, Xianghe, Tian, Kailu, Huang, Yan, Yang, Xi, Sun, Wen, Wang, Dong, Tsai, Yu-Hsuan, Luo, Tuoping, Li, Gang

Issue&Volume: 2024-06-04

Abstract: Recent advances in chemical proteomics have focused on developing chemical probes that react with nucleophilic amino acid residues. Although histidine is an attractive candidate due to its importance in enzymatic catalysis, metal binding and protein–protein interaction, its moderate nucleophilicity poses challenges. Its modification is frequently influenced by cysteine and lysine, which results in poor selectivity and narrow proteome coverage. Here we report a singlet oxygen and chemical probe relay labelling method that achieves high selectivity towards histidine. Libraries of small-molecule photosensitizers and chemical probes were screened to optimize histidine labelling, enabling histidine profiling in live cells with around 7,200 unique sites. Using NMR spectroscopy and X-ray crystallography, we characterized the reaction mechanism and the structures of the resulting products. We then applied this method to discover unannotated histidine sites key to enzymatic activity and metal binding in select metalloproteins. This method also revealed the accessibility change of histidine mediated by protein–protein interaction that influences select protein subcellular localization, underscoring its capability in discovering functional histidines.

DOI: 10.1038/s41557-024-01545-6

Source: https://www.nature.com/articles/s41557-024-01545-6