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Adeoye, RI, Babbudas, N, Birchenough, M, Giuntini, F 
ORCID: 0000-0002-3444-8183 and Olorunniji, FJ ORCID: 0000-0001-9389-2981
(2026)
Rational redesign of high-activity G-quadruplex DNAzyme through flanking and looping of nucleobases.
Scientific reports.
ISSN 2045-2322
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Rational redesign of high-activity G-quadruplex DNAzyme through flanking and looping of nucleobases..pdf - Published Version Available under License Creative Commons Attribution Non-commercial No Derivatives. Download (17MB) | Preview |
Abstract
G-quadruplex (G4) DNAzymes are guanine-rich oligonucleotides with intrinsic peroxidase-mimicking activity upon complexation with hemin, offering a promising alternative to protein-based enzymes in biosensing. However, their relatively low catalytic efficiency limits practical applications. Here, we present a structure-guided redesign of the high-activity aptamer [B7]-3-0 by incorporating strategic flanking and looping nucleobase modifications. Introduction of adenine and thymine–cytosine elements at the 3′ end led to up to 4-fold enhancements in reaction extent and a 3-fold increase in initial velocity under moderate hydrogen peroxide conditions (0.425 mM). Remarkably, the modified B730 variants retained activity at elevated H₂O₂ concentrations (4.25 mM), achieving up to 8-fold catalytic enhancement and outperforming high-activity DNAzymes including AS1411 and CatG4. These redesigned DNAzymes demonstrated improved peroxidase activity and resistance to oxidative inactivation, addressing a major limitation of both natural and artificial peroxidases. Our findings establish flanking and loop engineering as a cost-effective and broadly applicable strategy for optimizing G4 DNAzymes and underscore their potential in the development of next-generation biosensors.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | DNAzymes; G-quadruplex; Peroxidase; Rate-enhancement; 3101 Biochemistry and Cell Biology; 31 Biological Sciences |
| Subjects: | R Medicine > RS Pharmacy and materia medica |
| Divisions: | Pharmacy and Biomolecular Sciences |
| Publisher: | Springer Nature |
| Date of acceptance: | 7 January 2026 |
| Date of first compliant Open Access: | 22 January 2026 |
| Date Deposited: | 22 Jan 2026 09:28 |
| Last Modified: | 22 Jan 2026 09:28 |
| DOI or ID number: | 10.1038/s41598-026-35686-0 |
| URI: | https://researchonline.ljmu.ac.uk/id/eprint/27948 |
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