Hydrogel nanoparticle as an artificial nuclease for selective DNA cleavage

Controlled and sequence-selective DNA cleavage plays a crucial role in biomolecular engineering and nucleic acid technologies. Here, we report the development of cerium-coordinated catalytic hydrogel nanoparticles (Ce-CHNP) that exhibit single-strand-selective nuclease activity. Ce-CHNP was synthesized via a one-step polymerization process in which Ce(IV) coordinated with N-hydroxymethyl acrylamide to form nanoceria inside the polymer network. The artificial enzyme exhibits high efficiency for hydrolysis of phosphodiester, with kinetic parameters of Km = 1.20 mM, kcat = 13.57 s−1, and kcat/Km = 1.13 × 104 M−1 s−1 when evaluated using bis(4-nitrophenyl) phosphate as a model substrate. Ce-CHNP also demonstrates excellent stability across a wide pH range and at elevated temperatures. Mechanistic studies, including X-ray photoelectron spectroscopy (XPS), redox assays, and 31P solid-state NMR, elucidated the catalytic mechanism by which Ce(III) and Ce(IV) on nanoceria synergistically hydrolyze phosphodiester bonds. Notably, Ce-CHNP selectively hydrolyzes single-stranded DNA (ssDNA) in the presence of double-stranded DNA (dsDNA). For targeting double stranded DNA (dsDNA) sequences, peptide nucleic acid (PNA)-guided Ce-CHNP system functions as a programmable nuclease to cleave single strand regions generated by PNA invasion. Our results demonstrate that Ce-CHNP offers a programmable artificial nuclease platform useful for nucleic acid processing, and future biosensing, diagnostics and therapeutic applications.