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Updated: May 2, 2026
Quantitative, Real-time Analysis of Base Excision Repair Activity in Cell Lysates Utilizing Lesion-specific Molecular Beacons
Published on: August 6, 2012
A real-time biochemical assay for quantitative analyses of APOBEC-catalyzed DNA deamination
Christopher A Belica1,2,3, Michael A Carpenter4,5, Yanjun Chen4
1Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, Minnesota, USA, 55455.
Abstract:
Over the past decade, the connection between APOBEC3 cytosine deaminases and cancer mutagenesis has become increasingly apparent. This growing awareness has created a need for biochemical tools that can be used to identify and characterize potential inhibitors of this enzyme family. In response to this challenge, we have developed a Real-time APOBEC3-mediated DNA Deamination (RADD) assay. This assay offers a single-step set-up and real-time fluorescent read-out, and it is capable of providing insights into enzyme kinetics and also offering a high-sensitivity and easily scalable method for identifying APOBEC3 inhibitors. This assay serves as a crucial addition to the existing APOBEC3 biochemical and cellular toolkit and possesses the versatility to be readily adapted into a high-throughput format for inhibitor discovery.
Insights
Researchers developed a new assay to detect APOBEC3-mediated DNA deamination. This tool aids in discovering inhibitors for cancer mutagenesis, offering a sensitive and scalable method.
Area of Science:
- Biochemistry
- Molecular Biology
- Cancer Research
Background:
- The role of APOBEC3 cytosine deaminases in cancer mutagenesis is increasingly recognized.
- There is a need for biochemical tools to identify and characterize APOBEC3 inhibitors.
Approach:
- Developed a Real-time APOBEC3-mediated DNA Deamination (RADD) assay.
- The RADD assay features a single-step setup and real-time fluorescent readout.
- This method provides insights into enzyme kinetics and inhibitor identification.
Key Points:
- The RADD assay is highly sensitive and easily scalable.
- It is adaptable for high-throughput screening of potential APOBEC3 inhibitors.
- This assay complements existing APOBEC3 research tools.
Conclusions:
- The RADD assay is a valuable addition to the APOBEC3 research toolkit.
- It facilitates the discovery of novel inhibitors targeting cancer mutagenesis.
- The assay's versatility supports its adaptation for drug discovery pipelines.
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