Synthesis and Incorporation of a pH-Responsive Nucleoside Into DNA Sequences

DNA's programable thermodynamics, structural versatility, and ease of synthesis makes it an ideal material for constructing molecular devices. While many biological systems are powered by proton gradients to drive dynamic processes, harnessing pH differences in DNA nanotechnology is possible through pH-responsive DNA motifs. Existing strategies, however, often depend on strict sequence constraints or nonphysiological pH conditions, limiting their applicability in complex DNA origami structures. In this article, a nucleoside with pH-sensitive base pairing is developed that reversibly switches its pairing specificity near physiological pH. This unnatural building block is recognized by standard polymerases, and its pairing behavior can be controlled by pH. Characterization of the base pairing properties reveals that duplex stability varies with pH, while canonical sequences remain unaffected. This design enables programable sequence motifs that transition between duplex and single-stranded DNA in response to pH changes. Our unnatural nucleoside therefore provides a versatile tool for dynamic DNA nanotechnology, with potential applications in DNA nanomachines, biosensing, and targeted drug delivery. Additionally, its physiological pK_a may enable general acid-base catalysis in ribozymes or DNAzymes, analogous to histidine in protein enzymes.