Enzymes containing 3′→5′exonuclease activities play an important role in various key cellular and physiological processes.The development of fluorescence biosensor is an efficient method to detecting enzyme activit...Enzymes containing 3′→5′exonuclease activities play an important role in various key cellular and physiological processes.The development of fluorescence biosensor is an efficient method to detecting enzyme activity.Herein,a fluorescence resonance energy transfer(FRET)“on”and“off”strategy for detecting exonuclease III(Exo III)activity has been developed.We report here that the double-stranded DNA(dsDNA)enables to bind tightly to self-assembled nanosheets of cationic perylene monoimide derivative(PMI-O7)through electrostatic interaction,and the 6-carboxyfluorescein(FAM)-modified dsDNA could be efficiently quenched via FRET between FAM and PMI-O7.Upon the addition of Exo III,the dsDNA will be digested and the FAM fluorophore will be released,resulting in the fluorescence recovery of FAM.This method provides a simple and sensitive biosensor platform with a low detection limit of 0.077 U/mL for Exo III.Importantly,this method exhibits similar and calibration curves for the detection of Exo III in both buffer and fetal bovine serum samples,indicating that this platform has potential to detect Exo III activity in complex samples.展开更多
基金supported by the Natural Science Foundation of Guangdong Province,China (No.2014A030313194)the Science and Technology Program of Guangzhou,China (No.201804010017)+1 种基金the Science and Technology Program of Guangdong Province,China (No.2017A050506021)the National Natural Science Foundation of China (No.21374137).
文摘Enzymes containing 3′→5′exonuclease activities play an important role in various key cellular and physiological processes.The development of fluorescence biosensor is an efficient method to detecting enzyme activity.Herein,a fluorescence resonance energy transfer(FRET)“on”and“off”strategy for detecting exonuclease III(Exo III)activity has been developed.We report here that the double-stranded DNA(dsDNA)enables to bind tightly to self-assembled nanosheets of cationic perylene monoimide derivative(PMI-O7)through electrostatic interaction,and the 6-carboxyfluorescein(FAM)-modified dsDNA could be efficiently quenched via FRET between FAM and PMI-O7.Upon the addition of Exo III,the dsDNA will be digested and the FAM fluorophore will be released,resulting in the fluorescence recovery of FAM.This method provides a simple and sensitive biosensor platform with a low detection limit of 0.077 U/mL for Exo III.Importantly,this method exhibits similar and calibration curves for the detection of Exo III in both buffer and fetal bovine serum samples,indicating that this platform has potential to detect Exo III activity in complex samples.
