Chemoproteomic profiling of kinases in live cells using electrophilic sulfonyl triazole probes,Chemical Science

当前位置： X-MOL 学术 › Chem. Sci. › 论文详情

Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)

Chemoproteomic profiling of kinases in live cells using electrophilic sulfonyl triazole probes
Chemical Science ( IF 7.6 ) Pub Date : 2021-1-21 , DOI: 10.1039/d0sc06623k
Tao Huang ₁ , Seyyedmohsen Hosseinibarkooie ₂ , Adam L Borne ₃ , Mitchell E Granade ₃ , Jeffrey W Brulet ₁ , Thurl E Harris ₃ , Heather A Ferris ₂ , Ku-Lung Hsu _{1,

3,

4,

5}

Affiliation

Sulfonyl-triazoles are a new class of electrophiles that mediate covalent reaction with tyrosine residues on proteins through sulfur-triazole exchange (SuTEx) chemistry. Recent studies demonstrate the broad utility and tunability of SuTEx chemistry for chemical proteomics and protein ligand discovery. Here, we present a strategy for mapping protein interaction networks of structurally complex binding elements using functionalized SuTEx probes. We show that the triazole leaving group (LG) can serve as a releasable linker for embedding hydrophobic fragments to direct molecular recognition while permitting efficient proteome-wide identification of binding sites in live cells. We synthesized a series of SuTEx probes functionalized with a lipid kinase fragment binder for discovery of ligandable tyrosines residing in catalytic and regulatory domains of protein and metabolic kinases in live cells. We performed competition studies with kinase inhibitors and substrates to demonstrate that probe binding is occurring in an activity-dependent manner. Our functional studies led to discovery of probe-modified sites within the C2 domain that were important for downregulation of protein kinase C-alpha in response to phorbol ester activation. Our proof of concept studies highlight the triazole LG of SuTEx probes as a traceless linker for locating protein binding sites targeted by complex recognition elements in live cells.

中文翻译：

使用亲电磺酰三唑探针对活细胞中的激酶进行化学蛋白质组学分析

磺酰三唑是一类新型亲电子试剂，可通过硫三唑交换 (SuTEx) 化学介导与蛋白质上酪氨酸残基的共价反应。最近的研究证明了 SuTEx 化学在化学蛋白质组学和蛋白质配体发现方面的广泛实用性和可调性。在这里，我们提出了一种使用功能化 SuTEx 探针绘制结构复杂结合元件的蛋白质相互作用网络的策略。我们表明，三唑离去基团（LG）可以作为可释放的连接体，用于嵌入疏水性片段以指导分子识别，同时允许在活细胞中有效地在蛋白质组范围内识别结合位点。我们合成了一系列用脂质激酶片段结合剂功能化的 SuTEx 探针，用于发现活细胞中蛋白质和代谢激酶的催化和调节域中的可配体酪氨酸。我们用激酶抑制剂和底物进行了竞争研究，以证明探针结合是以活性依赖性方式发生的。我们的功能研究发现了 C2 结构域内的探针修饰位点，这些位点对于响应佛波酯激活而下调蛋白激酶 C-α 非常重要。我们的概念验证研究强调 SuTEx 探针的三唑 LG 作为无痕接头，用于定位活细胞中复杂识别元件所针对的蛋白质结合位点。

更新日期：2021-01-21

点击分享查看原文

点击收藏

公开下载

阅读更多本刊最新论文本刊介绍/投稿指南11