Precise Mechanochemical Scission of DNA Guided by Secondary Structures

DNA has recently been exploited as a biopolymer, enabling sequence-specific, mechanophore-free mechanochemistry with few-nucleotide-precision scission and monomer-level analysis of breakage. Strategically positioned nicks in double-stranded DNA (dsDNA) have previously been shown to act as mechanophores, concentrating force on the non-nicked strand opposite each nick to induce localized scission; however, base fraying inherently limits this precision. To address this, we identify, for the first time, the potential of DNA hairpins to serve as 'biomechanophores' and direct precise mechanochemical scission of DNA. Using a dsDNA construct containing a centrally embedded hairpin motif, we show that ultrasonication induces cleavage regioselectively opposite the hairpin loop. Force localization around nucleotides on the nonhairpin strand was rationalized by all-atom, constant-force molecular dynamics (MD) simulations. Next-generation sequencing (NGS) of sonicated fragments revealed a narrow, well-defined scission distribution on the nonhairpin strand, in contrast to a much broader, bimodal-like distribution on the hairpin strand. Together with the simulations, these results support a two-step cleavage mechanism. Our findings establish DNA hairpins as the first functional biomechanophores capable of directing mechanically induced strand scission with high spatial precision, expanding the mechanochemical toolbox for nucleic acid manipulation.