New Cornerstone Science Laboratory, Synthetic and Functional Biomolecules Center, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
Zhejiang Provincial Key Laboratory for Cancer Molecular Cell Biology, Life Sciences Institute, Zhejiang University, Hangzhou, China.
Nat Chem. 2024 Apr;16(4):533-542. doi: 10.1038/s41557-024-01463-7. Epub 2024 Feb 28.
Tryptophan (Trp) plays a critical role in the regulation of protein structure, interactions and functions through its π system and indole N-H group. A generalizable method for blocking and rescuing Trp interactions would enable the gain-of-function manipulation of various Trp-containing proteins in vivo, but generating such a platform remains challenging. Here we develop a genetically encoded N-vinyl-caged Trp capable of rapid and bioorthogonal decaging through an optimized inverse electron-demand Diels-Alder reaction, allowing site-specific activation of Trp on a protein of interest in living cells. This chemical activation of a genetically encoded caged-tryptophan (Trp-CAGE) strategy enables precise activation of the Trp of interest underlying diverse important molecular interactions. We demonstrate the utility of Trp-CAGE across various protein families, such as catalase-peroxidases and kinases, as translation initiators and posttranslational modification readers, allowing the modulation of epigenetic signalling in a temporally controlled manner. Coupled with computer-aided prediction, our strategy paves the way for bioorthogonal Trp activation on more than 28,000 candidate proteins within their native cellular settings.
色氨酸（Trp）通过其π系统和吲哚 N-H 基团在调节蛋白质结构、相互作用和功能方面发挥着关键作用。一种可普遍用于阻断和挽救 Trp 相互作用的方法将能够在体内对各种含有 Trp 的蛋白质进行功能获得性操作，但生成这样的平台仍然具有挑战性。在这里，我们开发了一种遗传编码的 N-乙烯基笼形色氨酸（N-vinyl-caged Trp），通过优化的逆电子需求 Diels-Alder 反应能够快速进行生物正交脱笼，从而能够在活细胞中对感兴趣的蛋白质进行位点特异性的 Trp 激活。这种遗传编码的笼形色氨酸（Trp-CAGE）策略的化学激活能够精确地激活各种重要分子相互作用下的感兴趣的 Trp。我们证明了 Trp-CAGE 在各种蛋白质家族中的用途，如过氧化氢酶过氧化物酶和激酶，作为翻译起始子和翻译后修饰阅读器，允许以时间控制的方式调节表观遗传信号。结合计算机辅助预测，我们的策略为在其天然细胞环境中的超过 28000 个候选蛋白质上进行生物正交 Trp 激活铺平了道路。
