The pace of progress in biomedical research directly depends on techniques that enable the quantitative interrogation of interactions between proteins and other biopolymers, or with their small-molecule ligands. Time-resolved Förster resonance energy transfer (TR-FRET) assay platforms offer high sensitivity and specificity. However, the paucity of accessible and biocompatible luminescent lanthanide complexes, which are essential reagents for TR-FRET-based approaches, and their poor cellular permeability have limited broader adaptation of TR-FRET beyond homogeneous and extracellular assay applications. Here, we report the development of CoraFluors, a new class of macrotricyclic terbium complexes, which are synthetically readily accessible, stable in biological media and exhibit photophysical and physicochemical properties that are desirable for biological studies. We validate the performance of CoraFluors in cell-free systems, identify cell-permeable analogs and demonstrate their utility in the quantitative domain-selective characterization of Keap1 ligands, as well as in isoform-selective target engagement profiling of HDAC1 inhibitors in live cells.

生物医学研究的进展速度直接取决于能够定量研究蛋白质与其他生物聚合物或其小分子配体之间相互作用的技术。时间分辨福斯特共振能量转移 (TR-FRET) 检测平台具有高灵敏度和特异性。然而，由于缺乏可利用且生物相容的发光稀土配合物（这些配合物是基于 TR-FRET 的方法的重要试剂），而且它们的细胞渗透性差，限制了 TR-FRET 在同质和细胞外测定应用之外的更广泛的适应性。在此，我们报告了 CoraFluors 的开发，这是一种新型大三环铽配合物，其易于合成，在生物介质中稳定，并表现出生物学研究所需的光物理和物理化学性质。我们验证了 CoraFluors 在无细胞系统中的性能，鉴定了细胞可渗透的类似物，并证明了它们在 Keap1 配体的定量域选择性表征以及活细胞中 HDAC1 抑制剂的异构体选择性靶点接合分析中的实用性。