Development of a Nitric Oxide-Responsive Labeling Reagent for Proteome Analysis of Live Cells
Development of a Nitric Oxide-Responsive Labeling Reagent for Proteome Analysis of Live Cells
复制标题
开发用于活细胞蛋白质组分析的一氧化氮响应标记试剂
DOI：
10.1021/acschembio.8b01021
复制
发表时间：
2019
影响因子：
4
通讯作者：
Hamachi Itaru
中科院分区：
文献类型：
--
作者：
Nishikawa Yuki;Miki Takayuki;Awa Masashi;Kuwata Keiko;Tamura Tomonori;Hamachi Itaru
一氧化氮（NO）是一种多效性信号分子，参与心血管、神经和免疫系统的多种生理和病理生理机制的调节。为了详细了解NO的生物学功能，在高NO浓度环境中发现的蛋白质的全面表征是至关重要的。在此，我们描述了基于N-烷氧基酰基邻苯二胺作为最佳反应支架的NO响应蛋白标记试剂的设计。所设计的分子可以标记蛋白质在小鼠巨噬细胞中响应内源性产生的NO。NO响应蛋白质标记和液相色谱-串联质谱技术的组合允许NO产生的条件下的蛋白质组的表征。此外，我们证明了我们的试剂能够选择性地标记并用于荧光可视化混合细胞培养系统中的NO产生细胞。
Nitric oxide (NO) is a pleiotropic signaling molecule involved in the regulation of diverse physiological and pathophysiological mechanisms in cardiovascular, nervous, and immunological systems. To understand the biological functions of NO in detail, comprehensive characterization of proteins found in high-NO concentration environments is crucial. Herein, we describe the design of NO-responsive protein labeling reagents based onN-alkoxyacyl-o-phenylenediamine as an optimal reactive scaffold. The designed molecules can label proteins in murine macrophage cells in response to endogenously produced NO. The combination of NO-responsive protein labeling and liquid chromatography–tandem mass spectrometry technology allowed the characterization of the proteome under NO-generated conditions. Moreover, we demonstrated that our reagent was able to selectively mark and be used to fluorescently visualize NO-producing cells in a mixed cell culture system.