Modulating Aptamer Specificity with pH-Responsive DNA Bonds

Abstract

Aptamers that recognize specific cells in a complex environment have emerged as invaluable molecular tools in bioanalysis and in the development of targeted therapeutics. The selective recognition of aptamers, however, can be compromised by the coexistence of target receptors on both target cells and other cells. To address this problem, we constructed a structure-switchable aptamer (SW-Apt) with reconfigurable binding affinity in accordance with the microenvironment of target cells. The SW-Apt makes use of i-motifs, which are quadruplex structures that form in sequences rich in cytosine. More specifically, we report the design of single-stranded, pH-responsive i-motif-modified aptamers able to bind specifically with target cells by exploiting their pH. Here, the i-motif serves as a structural domain to either facilitate the binding ability of aptamers to target cells or suppress the binding ability of aptamers to nontarget cell based on the pH of the cellular microenvironment. SW-Apt exhibited high binding ability with target cells at acidic pH, while no obvious binding was observed at physiological pH. The i-motif-induced structure-switching was verified with Förster resonance energy transfer and circular dichroism spectroscopy. Notably, SW-Apt exhibits high specificity in serum and excellent stability, likely attributed to the folded quadruplex i-motif structure. This study provides a simple and efficient strategy to chemically modulate aptamer binding ability and thus improve aptamer binding specificity to target cells, irrespective of the coexistence of identical receptors on target and nontarget cells.

Rational design of aptamer switches with programmable pH response

“…The DeRosa group used noncanonical base pairing between guanine and protonated adenine to disrupt G-quadruplex formation in a thrombin aptamer, enabling reversible pH-mediated binding of thrombin at pH 7 followed by release at pH 5 15 . Most recently, the Tan group employed a cytosine-rich i-motif structure to preferentially stabilize folding of a tyrosine kinase-7 aptamer in acidic conditions, demonstrating selective binding to the aptamer's membrane-bound target at low pH 16 . However, all these approaches entail the use of defined DNA motifs that alter the aptamer binding structure under very particular pH conditions.…”

Engineering Aptamer Switches for Multifunctional Stimulus‐Responsive Nanosystems

“…For example, the Tan group recently reported a strategy to tune aptamer binding through the incorporation of the i‐motif ( Figure A). [ 51 ] Their design consisted of a recognition domain flanked by linker sequences and a split i‐motif structure that acted as a modulator. At neutral pH, the i‐motif will be unstructured, preventing hybridization of the linker domain and thereby impairing folding of the aptamer binding loop.…”

“…Adapted with permission. [ 51 ] Copyright 2018, American Chemical Society. B) Achieving pH‐induced allosteric control by introducing C–T triplexes at the 3′ end of a cocaine‐binding aptamer with a three‐way junction structure (top).…”

Engineering Cell‐Surface Receptors with DNA Nanotechnology for Cell Manipulation

“…Moreover, DNA sequences rich in guanine and cytosine can form stable quadruplexes in the presence of specific metal ions and low pH, and this can be reversibly controlled by changing those environmental parameters (Figure C, iv) . The DNA i‐motif structure, for example, was applied to achieve reversible control based on a conformation change at pH lower than 6.5 …”

“…[38] The DNA i-motif structure,f or example, was applied to achieve reversible control based on ac onformation change at pH lower than 6.5. [39] In addition, discrete 3D DNA origami nanostructures based on weak blunt-end stacking interaction between DNA helices with shape complementarity can be nanofabricated. These allow reversible reconfiguration by changingt he cation concentration or temperature( Figure 2C,v).…”