State Key Laboratory of Molecular Reaction Dynamics, Dalian Institute of Chemical Physics, Chinese Academy of Sciences (CAS), Dalian 116023, China.
Department of Pharmacy, Peking University Shenzhen Hospital, Shenzhen 518036, China.
J Med Chem. 2021 Nov 11;64(21):16177-16186. doi: 10.1021/acs.jmedchem.1c01469. Epub 2021 Oct 25.
In the need for improving the labeling quality of super-resolution imaging, multifarious fluorescent labeling strategies have sprang up. Among them, a small molecule inhibitor-probe (SMI-probe) shows its advancement in fine mapping due to its smaller size and its specific binding to a specific site. Herein, we report a novel protocol of mechanism-guided directional modification of fluorophores into fluorescent inhibitors for enzyme targeting, which could half the size of the SMI-probe. To confirm the feasibility of the strategy, carboxylesterase (hCE) inhibitors are designed and developed. Among the constructed molecule candidates, NIC-4 inhibited both isoforms of hCE1 and hCE2, with IC values of 4.56 and 4.11 μM. The CE-targeting specificity of NIC-4 was confirmed by colocalizing with an immunofluorescent probe in fixed-cell confocal imaging. Moreover, NIC-4 was used in live-cell super-resolution microscopy, which indicates dotlike structures instead of the larger staining with the immunofluorescent probe. Moreover, it enables the real-time tracking of dynamic flow of carboxylesterases in live cells.
为了提高超分辨率成像的标记质量，出现了各种荧光标记策略。其中，小分子抑制剂探针（SMI-probe）由于其较小的尺寸及其与特定位点的特异性结合，在精细作图方面显示出了其优势。在此，我们报告了一种将荧光团定向修饰为酶靶向荧光抑制剂的新方法，该方法可以将 SMI-probe 的尺寸缩小一半。为了验证该策略的可行性，我们设计并开发了羧酸酯酶（hCE）抑制剂。在构建的候选分子中，NIC-4 抑制了 hCE1 和 hCE2 的两种同工酶，IC 值分别为 4.56 和 4.11 μM。通过与固定细胞共聚焦成像中的免疫荧光探针共定位，证实了 NIC-4 的 CE 靶向特异性。此外，NIC-4 被用于活细胞超分辨率显微镜，其结果显示点状结构，而不是免疫荧光探针的更大染色。此外，它还能够实时跟踪活细胞中羧酸酯酶的动态流动。
