Acidic Extracellular\npH-Activated Allosteric DNA Nanodevice\nfor Fluorescence Imaging of APE1 Activity in Tumor Cells

Allostery is a phenomenon where the binding of a ligand\nat one\nallosteric site influences the affinity for another ligand at an active\nsite. Different from orthosteric regulation, it allows for more precise\ncontrol of biomolecular activity and enhances the stability of the\nmolecules. Inspired by allosteric regulation of natural molecules,\nwe present a Y-shaped allosteric DNA nanodevice, termed YssAP, that\nwas pH-responsive and functionalized with the AS1411 aptamer for accurate\nfluorescence imaging of human apurinic/apyrimidinic endonuclease (APE1)\nactivity in tumor cells. With rational design, YssAP could not be\ncut by APE1, and Cy5 was in the proximity of BHQ2, leading to suppressed\nsignal emission. In contrast, since acidic pH acted as an allosteric\neffector, YssAP underwent a conformational change into an activated\nDNA probe (YdsAP) at acidic extracellular pH. After entering the tumor\ncell via the specific recognition of AS1411 aptamer, the overexpressed\nAPE1 in the tumor cell cut the AP site on YdsAP. Cy5 moved far away\nfrom BHQ2, resulting in a strong signal output. Compared with the\ndirect construction of the APE1 substrate, allosteric DNA nanodevices\nhave more accurate imaging effects, which can be precisely adjusted\nby changing the switching state. We anticipate that this strategy\nwill be applied in the screening of APE1 inhibitors and precise tumor\ndiagnosis.