基于细胞透性活动的乌比基因探针可实现人体脱基因酶的细胞内分析
Weijun Gui1, Christine A Ott1, Kun Yang1
1Department of Chemistry and Biochemistry , University of Delaware , 214A Drake Hall , Newark , Delaware 19716 , United States.
Journal of the American Chemical Society
|September 22, 2018
概括
研究人员开发了基于细胞透性二维基酶活性的探针 (DUB ABPs) 用于细胞内分析. 这些探测器可以在完整的细胞中研究DUB,从而促进对二维基因酶 (DUB) 的理解和药物发现.
科学领域:
- 生物化学
- 细胞生物学
- 化学生物学
背景情况:
- 脱基因酶 (DUB) 是许多细胞过程中的关键酶.
- 现有的基于ubiquitin (Ub) 的DUB活动探针 (DUB ABPs) 往往缺乏细胞透性,仅限于细胞溶解物.
- 细胞溶解可以破坏细胞组织并改变DUB活动,阻碍生理相关的研究.
研究的目的:
- 开发新的细胞透性DUBABP,用于细胞内DUB活动的分析.
- 在完整的活细胞中研究DUB及其功能.
- 推进在细胞环境中对DUB的研究,并支持药物发现工作.
主要方法:
- 一种半合成方法被用来创建模块化,基于ubiquitin的DUB探测器与反应性弹头.
- 使用细胞透 (CPP),特别是循环聚氨酸 (cR10),以增强DUB ABPs的细胞内传递.
- 活细胞光显微镜证实了探针的输送;用于完整的HeLa细胞中的DUB概况的免疫捕获和质谱学.
主要成果:
- 一个新的细胞透性DUBABP类型已成功生成和验证.
- 与TAT相比,循环聚氨酸 (cR10) 显示了DUB ABPs的细胞内传递的增强.
- 在完整的HeLa细胞中进行细胞内DUB分析,确定了活性DUB,并使用探针评估局部抑制剂活性.
结论:
- 细胞透性DUB ABPs提供了一个强大的工具,用于在本地细胞环境中调查DUBs.
- 这项技术有助于更深入地了解DUB的功能和细胞通路.
- 开发的探测器对推进针对人类DUB的药物发现具有重大前景.
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