INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
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2025
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IF 8.5
Jingqi Sun,Jiaxin Li,Yanzhi Xing,Han Leng,Wei Chen,Yanfeng Zhang,Xuwei Chen
Adenine DNA glycosylase (MutY) is a crucial member of DNA glycosylase family, and the abnormal expression of the human MutY homologs is associated with the pathogenesis of various diseases, therefore, convenient and cost-effective assessing the activity of MutY holds significant biological and medical importance. Herein, the precise identification/excision capacity of MutY to mismatched G-A base pair of dsDNA chains and the DNA-template-dependant fluorescence behaviors of copper nano cluster (CuNCs) was exploited for the accurate sensing of active MutY. Hairpin DNA with G-A base mismatch was excised by MutY to produce dsDNA chains with repetitive AAT-TTA base pairs. The newly formed dsDNA provided more active sites for the growth of CuNCs compared to the original hairpin DNA, resulting in the significantly enhanced fluorescence of final CuNCs. MutY was accurately quantified with a detection limit of 9.98 nmol L −1 . The developed sensing protocol exhibited excellent selectivity toward MutY over various ions, neutral biomolecules, and protein species. Most importantly, The sensing system is capable to distinguish the active MutY from other MutY homologs with low activity, e.g., de-[4Fe4S] cluster MutY (DIS-MutY), and the practicality was well demonstrated by detecting active MutY contents in various cell lysates.
研究领域:
生物化学
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分子生物学
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医学诊断
✓ 已验证
产品关联
DOI
PubMed
高影响因子
ELECTROCHIMICA ACTA
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2015
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IF 4.504
Tingting Zhang,Huimin Zhao,Xie Quan,Shuo Chen
An electrochemiluminescence (ECL) sensing for sensitive assaying of active DNA glycosylase was developed using the enhanced host-guest recognition technique. The α-cyclodextrin was selected as the host molecule which captured the guest-labeled ECL probe to the surface of electrode by the host-guest recognition. The ECL probe can be protected from the digestion of exonuclease I (Exo I) and exonuclease III (Exo III) with the presence of the target enzyme human 8-oxoguanine DNA glycosylase 1 (hOGG 1), resulting in the ECL emission intensity was correlated with the quantity of hOGG 1. The α-cyclodextrin functionalized gold/silica (Au/SiO 2 ) cell-shell nanoparticles was prepared to enhance the host-guest recognition sensitivity. Because of the increased recognition sites provided by α-cyclodextrin functionalized Au/SiO 2 cell-shell nanoparticles, the nanoparticle-modified electrode displayed a high capacity for the guest ECL probe, and four fold enhancement of the ECL signals was achieved. The as-prepared ECL sensing exhibits excellent analytical property for the detection of hOGG 1 in the linear range of 2–100 U mL −1 with a detection limit of 0.225 U mL −1 (S/N = 3). Interference tests show that the ECL intensity of the interferents human apurinic/apyrimidinic endonuclease (APE 1), T4 endonuclease V (T4 PDG) and endonuclease III (Endo III) is within 36.5 to 44.7% of that of hOGG 1. The ECL sensing exhibits long-term stability with a relative standard deviation (RSD) of 1.6% for 16 cycles of continuous potential scans, and the life time of the ECL sensing is up to 20 days. The obtained results in this study indicate that the proposed ECL sensing possesses high sensitivity, specificity and stability and provides a powerful tool for assaying hOGG 1 activity.
研究领域:
生物化学
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分析化学
✓ 已验证
产品关联
DOI
高影响因子
MICROCHEMICAL JOURNAL
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2025
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IF 5.1
Ruyue Cong,Kaiyue Hu,Dandan Hu,Jingran Sun,Jinglu Yao,Zhiyong Guo,Sui Wang,Yufang Hu
The detection of UDG activity is essential for understanding DNA damage repair mechanisms, assessing cellular genetic stability, and advancing diagnostics and treatments for related diseases. An advanced electrochemical biosensor based on a silver-enhanced copper metal–organic framework (Ag@Cu-MOF) has been developed for the sensitive and selective detection of uracil-DNA glycosylase (UDG) activity in cellular environments. This design incorporates a prism substrate to stabilize the liquid phase on the electrode surface, with DNA probes immobilized on the prism surface. Upon UDG-catalyzed uracil excision, the probe undergoes structural changes, initiating a cascade reaction. The abundance of free DNA ends facilitates the approach of MOF materials to the electrode interface, creating a stable environment conducive to H 2 O 2 reduction, thereby amplifying the electrochemical signal. Comparative catalytic analyses revealed that the Ag@Cu-MOF-modified electrode, with its enhanced conductivity and high surface area, achieved efficient catalysis compared to Cu-MOF. The biosensor exhibited a linear response to UDG concentrations ranging from 0.00002 to 0.5 U/mL, with an impressively low detection limit of 0.00002 U/mL, highlighting its capability for quantitative analysis. The sensor demonstrated high selectivity, minimal interference from other biomolecules, and stable performance across multiple cycles, underscoring its robustness. Furthermore, UDG activity was successfully detected at varying cell densities and under hypoxic conditions, providing valuable insights into cellular DNA repair mechanisms and responses to stress. In summary, the Ag@Cu-MOF-based biosensor offers a promising platform for detecting UDG activity in cancer diagnostics and cellular research, with potential applications in monitoring DNA repair activities under both physiological and pathological conditions.
研究领域:
生物化学
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分子生物学
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分析化学
✓ 已验证
产品关联
DOI
高影响因子
TALANTA
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2018
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IF 4.244
Guirong Liu,Wenli He,Chenghui Liu
As an important base excision repair (BER) enzyme, uracil-DNA glycosylase (UDG) can repair the uracil-induced DNA lesion and maintain the genomic integrity. Herein, we have proposed a sensitive UDG assay based on the terminal deoxynucleotidyl transferase (TdT)-assisted formation of fluorescent copper nanoclusters (CuNCs). In this study, a uracil-containing stem-loop DNA substrate is rationally designed with its 3′-end blocked with 2′, 3′-dideoxycytosine (ddC). UDG can remove the uracil in the DNA substrate to generate an apurinic/apyrimidinic (AP) site, which can then be specifically cleaved by endonuclease IV (Endo IV) to expose a 3′-OH terminus. TdT will initiate the template-free DNA extension along the exposed 3′-OH terminus to produce a quite long poly(T) tail, which will perfectly template the production of fluorescent CuNCs. By recording the fluorescence of the CuNCs, the UDG activity can be faithfully detected. In contrast, if UDG is absent, the 3′-ddC terminus of the DNA substrate cannot be recognized by TdT and thus no TdT-based extension and formation of CuNCs will occur. The use of ddC as a 3′-end blocker can greatly decrease the nonspecific DNA extension and improve the signal-to-noise ratio. Furthermore, TdT is a template-free and sequence-independent DNA polymerase, which can effectively catalyze the tailing process up to a maximum of thousands of thymines , and each tail can form many fluorescent CuNCs. Therefore, an ultrahigh sensitivity is achieved and as low as 0.00005 U/mL of UDG can be clearly detected. Moreover, with an additional AP site-contained poly(A) oligonucleotide during TdT-mediated extension, a branched amplification mechanism is also preliminarily devised, which can further push the detection limit of UDG to an extremely low level of 0.000002 U/mL.
研究领域:
生物化学
·
分子生物学
·
分析化学
✓ 已验证
DOI
PubMed
高影响因子
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