Key Laboratory of Environmental Medicine Engineering of Ministry of Education, School of Public Health, Southeast University, Nanjing 210009, PR China.
Jiangsu Engineering Laboratory of Smart Carbon-Rich Materials and Device, School of Chemistry and Chemical Engineering, Southeast University, Nanjing 210009, PR China.
Anal Chem. 2021 Oct 12;93(40):13687-13693. doi: 10.1021/acs.analchem.1c03332. Epub 2021 Sep 29.
Uracil DNA glycosylase (UDG) is one of the key initiators for the base excision repair pathway. Since abnormal UDG expression is associated with various diseases, sensitive detection of UDG activity is critical for early clinical diagnosis. Here, a smart catalyzed hairpin assembly (CHA)-DNAzyme nanosystem is developed for intracellular UDG imaging by incorporating CHA and DNAzyme onto MnO nanosheets. In this strategy, the biodegradable MnO nanosheets are employed as nanocarriers for efficiently adsorbing and delivering five DNA probes into cells by endocytosis. Then, the MnO nanosheets are degraded by cellular glutathione to release the DNA modules at the same intracellular position. Liberated Mn, an indispensable DNAzyme cofactor, was used to promote catalytic cleavage for facilitating the cascade process in cells. Based on the uracil site-recognition and -excision operation of the target UDG, the activated CHA-DNAzyme nanosystem generates lots of DNAzyme-assisted CHA products, turning on the fluorescence resonance energy transfer response. This autocatalytic CHA-DNAzyme nanosystem provides a detectable minimum UDG concentration of 0.23 mU/mL, which is comparable to some reported UDG detection approaches. As a multiple signal amplification strategy, the CHA-DNAzyme nanosystem realizes the UDG imaging in living cells with enhanced sensitivity, indicating great promise in the prediction and diagnosis of early-stage cancer.
尿嘧啶 DNA 糖基化酶（UDG）是碱基切除修复途径的关键启动子之一。由于异常的 UDG 表达与各种疾病有关，因此敏感检测 UDG 活性对于早期临床诊断至关重要。在这里，通过将 CHA 和 DNA 酶整合到 MnO 纳米片上，开发了一种智能催化发夹组装（CHA）-DNA 酶纳米系统，用于通过细胞内 UDG 成像。在该策略中，可生物降解的 MnO 纳米片被用作纳米载体，通过内吞作用将五个 DNA 探针高效地吸附并递送到细胞中。然后，细胞内的谷胱甘肽将 MnO 纳米片降解，在相同的细胞内位置释放 DNA 模块。释放的 Mn 是 DNA 酶必不可少的辅助因子，用于促进细胞内级联过程中的催化裂解。基于目标 UDG 的尿嘧啶位点识别和切除操作，激活的 CHA-DNA 酶纳米系统产生大量 DNA 酶辅助的 CHA 产物，开启荧光共振能量转移反应。这种自催化的 CHA-DNA 酶纳米系统提供了可检测的最低 UDG 浓度为 0.23 mU/mL，可与一些报道的 UDG 检测方法相媲美。作为一种多重信号放大策略，CHA-DNA 酶纳米系统实现了活细胞中的 UDG 成像，具有增强的灵敏度，有望用于早期癌症的预测和诊断。
