Beijing National Laboratory for Molecular Sciences, Key Laboratory of Analytical Chemistry for Living Biosystems, Institute of Chemistry, Chinese Academy of Sciences, Beijing, 100190, China.
Huzhou Key Laboratory of Medical and Environmental Applications Technologies, School of Life Sciences, Huzhou University, Zhejiang, 313000, China.
Angew Chem Int Ed Engl. 2022 Aug 15;61(33):e202205043. doi: 10.1002/anie.202205043. Epub 2022 Jul 5.
Many fluorophores/probes suffer from the interference of albumin in biosystems. Herein, we propose an effective strategy to overcome this interference by virtue of both an albumin-insensitive fluorophore and its changeable π-conjugation, and demonstrate the strategy by designing an oxazine-based fluorogenic probe for aminopeptidase N (APN). The modification on the N atom in the oxazine fluorophore with alanine through a cleavable linker locks the resulting probe in a non-conjugated, colorless and non-fluorescent state, so the non-specific interaction of albumin produces no spectroscopic response. APN can selectively cleave the alanine moiety, restoring the large π-conjugation and strong fluorescence. The capability of the probe to eliminate the albumin influence has been demonstrated by imaging APN in different cell lines, and by quantitatively determining APN in human serum and mouse urine. The present strategy may be useful for developing more specific fluorogenic probes for other enzymes.
许多荧光团/探针受到生物体系中白蛋白的干扰。在此，我们提出了一种有效的策略，通过利用一种对白蛋白不敏感的荧光团及其可变化的π共轭，来克服这种干扰，并通过设计一种基于恶嗪的氨肽酶 N（APN）荧光探针来证明该策略。通过在恶嗪荧光团的 N 原子上用丙氨酸通过可裂解的连接子进行修饰，将所得探针锁定在非共轭、无色和非荧光状态，因此白蛋白的非特异性相互作用不会产生光谱响应。APN 可以选择性地切割丙氨酸部分，恢复大的π共轭和强荧光。该探针消除白蛋白影响的能力已通过在不同细胞系中成像 APN 以及定量测定人血清和小鼠尿液中的 APN 得到证明。该策略可能有助于开发其他酶的更特异的荧光探针。
