LAUSR

For proteins and DNA secondary structures such as G-quadruplexes and i-motifs, nanoconfinement can facilitate their folding and increase structural stabilities. However, properties of physiologically prevalent B-DNA duplex have not been elucidated inside nanocavity. By using a 17-bp DNA duplex in the form of a hairpin stem, here, we probed folding and unfolding transitions of the hairpin DNA duplex inside a DNA origami nanocavity. Compared to the free solution, the DNA hairpin inside the nanocage with a 15×15 nm cross section showed a drastic decrease in mechanical (20→9 pN) and thermodynamic (25→6 kcal/mol) stabilities. Free energy profiles revealed that activation energy of unzipping the hairpin DNA duplex decreased dramatically (28→8 kcal/mol) whereas the transition state moved closer to the unfolded state inside nanocage. All these indicate that nanoconfinement weakens the stability of hairpin DNA duplex to an unexpected extent. In a DNA hairpin made of a stem that contains complementary telomeric G-quadruplex (GQ) and i-motif (iM) forming sequences, the formation of the Hoogsteen base pairs underlining the GQ or iM is preferred over the Watson-Crick base pairs in the DNA hairpin. These results shed light on the behavior of DNA in nanochannels, nanopores, or nanopockets of various natural or synthetic machineries. It also elucidates an alternative pathway to populate non-canonical DNA over B-DNA in cellular environment where nanocavity is abundant.