Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory of Chemo/Bio-Sensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Aptamer Engineering Center of Hunan Province , Hunan University , Changsha 410082 , China.
Institute of Molecular Medicine (IMM), Renji Hospital, Shanghai Jiao Tong University School of Medicine, and College of Chemistry and Chemical Engineering , Shanghai Jiao Tong University , Shanghai 200240 , China.
ACS Appl Mater Interfaces. 2019 May 15;11(19):17722-17729. doi: 10.1021/acsami.9b04555. Epub 2019 May 6.
Selenocysteine (Sec) has proven to be the dominant active site of diverse selenoproteins that are directly linked with human health and disease. Thus, understanding the critical functions and dynamics of endogenous Sec at cellular and tissue levels is highly demanded. However, no method has been reported that is capable of providing reliable quantitative imaging analysis of Sec in living systems, especially in deep tissues, with low background signal and high sensitivity and imaging resolution simultaneously. To address this challenge, we herein report a novel class of engineered Sec-responsive fluorescent nanoprobes that combines two-photon excitation with Förster resonance energy transfer (FRET) mechanisms for direct, yet selective, sensing and imaging of biological Sec over abundant competing biothiols. Specifically, the two-photon excitation at the near-infrared window can minimize light scattering and background signals in tissues, thus offering improved spatial and temporal imaging of deep living tissues with reduced background interference. Moreover, a reasonable FRET donor-acceptor pair has further been designed and verified by theoretical calculation. The acceptor undergoes intramolecular rearrangement specifically in response to the nucleophilic attack of Sec, hence triggering remarkable FRET-mediated ratiometric fluorescence enhancement for sensitive and reliable quantification of Sec through self-calibration of two emission channels. These striking properties, along with good water solubility and biocompatibility, suggest that this strategy may serve as a valuable imaging tool for studying various Sec-related biological events in complex biological systems.
硒代半胱氨酸（Sec）已被证明是多种与人类健康和疾病直接相关的硒蛋白的活性中心。因此，在细胞和组织水平上深入了解内源性 Sec 的关键功能和动态具有重要意义。然而，目前还没有报道能够提供可靠的定量成像分析方法，用于在活体系统中，特别是在深部组织中，同时具有低背景信号、高灵敏度和高成像分辨率的 Sec。为了解决这一挑战，我们在此报告了一类新型的工程化 Sec 响应荧光纳米探针，它结合了双光子激发和Förster 共振能量转移（FRET）机制，用于直接但选择性地检测和成像生物体内丰富的竞争生物硫醇中的 Sec。具体而言，近红外窗口的双光子激发可以最大限度地减少组织中的光散射和背景信号，从而提供对深部活体组织的改进的时空成像，减少背景干扰。此外，还通过理论计算进一步设计和验证了合理的 FRET 供体-受体对。受体在 Sec 的亲核攻击下发生分子内重排，从而触发显著的 FRET 介导的比率荧光增强，通过两个发射通道的自校准实现 Sec 的敏感和可靠定量。这些显著的特性，加上良好的水溶性和生物相容性，表明该策略可能成为研究复杂生物系统中各种与 Sec 相关的生物事件的有价值的成像工具。
