Target-Activated DNA Polymerase Activity for Sensitive RNase H Activity Assay

Biotechnology Journal
Yujin JungHyun Gyu Park

Abstract

Herein, the ribonuclease H (RNase H) activity assay based on the target-activated DNA polymerase activity is described. In this method, a detection probe composed of two functional sequences, a binding site for DNA polymerase and a catalytic substrate for RNase H, serves as a key component. The detection probe, at its initial state, suppresses the DNA polymerase activity, but it becomes destabilized by RNase H, which specifically hydrolyzes RNA in RNA/DNA hybrid duplexes. As a result, DNA polymerase recovers its activity and initiates multiple primer extension reactions in a separate TaqMan probe-based signal transduction module, leading to a significantly enhanced fluorescence "turn-on" signal. This assay can detect RNase H activity as low as 0.016 U mL-1 under optimized conditions. Furthermore, its potential use for evaluating RNase H inhibitors, which have been considered potential therapeutic agents against acquired immune deficiency syndrome (AIDS), is successfully explored. In summary, this approach is quite promising for the sensitive and accurate determination of enzyme activity and inhibitor screening.

References

Jan 1, 1990·Pharmacology & Therapeutics·U Wintersberger
Jan 1, 1984·Molecular & General Genetics : MGG·T Ogawa, T Okazaki
Sep 25, 1999·The Journal of Biological Chemistry·H WuS T Crooke
Mar 29, 2001·Protein Science : a Publication of the Protein Society·A MuroyaK Morikawa
Jul 6, 2001·Journal of the American Chemical Society·J RenJ B Chaires
Sep 25, 2002·Molecular and Cellular Probes·J RizzoP Lu
Jan 22, 2003·Transplant Infectious Disease : an Official Journal of the Transplantation Society·G WuS Kusne
Jul 22, 2004·Analytical Biochemistry·King C ChanJohn A Beutler
Jan 9, 2008·Chembiochem : a European Journal of Chemical Biology·Yan ChenWeihong Tan
Feb 12, 2008·Virus Research·Sharon J Schultz, James J Champoux
May 12, 2015·Chemical Communications : Chem Comm·Ki Soo ParkHyun Gyu Park
Feb 26, 2016·Chemical Communications : Chem Comm·Ki Soo ParkHyun Gyu Park
Nov 26, 2016·Biosensors & Bioelectronics·Chuan ZhaoBin Liu

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Citations

Jul 10, 2020·Nucleic Acids Research·Yingxin HuLinqiang Pan
Apr 27, 2021·Journal of Materials Chemistry. B, Materials for Biology and Medicine·Arun Richard Chandrasekaran
Aug 20, 2021·Biotechnology Journal·Kingsley L Wong, Juewen Liu

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