Abstract
Aptamer switches that respond sensitively to pH could enhance control over molecular devices, improving their diagnostic and therapeutic efficacy. Previous designs have inserted pH-sensitive DNA motifs into aptamer sequences. Unfortunately, their performance was limited by the motifs’ intrinsic pH-responses and could not be tuned to operate across arbitrary pH ranges. Here, we present a methodology for converting virtually any aptamer into a molecular switch with pH-selective binding properties — in acidic, neutral, or alkaline conditions. Our design inserts two orthogonal motifs that can be manipulated in parallel to tune pH-sensitivity without altering the aptamer sequence itself. From a single ATP aptamer, we engineer pH-controlled target binding under diverse conditions, achieving pH-induced selectivity in affinity of up to 1,000-fold. Importantly, we demonstrate the design of tightly regulated aptamers with strong target affinity over only a narrow pH range. Our approach offers a highly generalizable strategy for integrating pH-responsiveness into molecular devices.
Highlights
Aptamer switches that respond sensitively to pH could enhance control over molecular devices, improving their diagnostic and therapeutic efficacy
Physiological pH conditions are maintained through tight homeostatic control within cells and tissues, and local variations in pH play a critical role in a number of important biological processes[1,2]
We have generated an array of pH-responsive strand displacement (PSD) aptamer switches, all based on the same core aptamer sequence, that have been engineered to preferentially bind or release their molecular target under various pH conditions
Summary
Aptamer switches that respond sensitively to pH could enhance control over molecular devices, improving their diagnostic and therapeutic efficacy. We describe here a general strategy that should make it possible to convert virtually any aptamer into a pH-responsive switch that can be tuned to selectively undergo a dramatic shift in affinity under acidic, neutral, or alkaline conditions We achieve this pH specificity by introducing two orthogonal modes of control that can be manipulated in parallel to tune sensitivity to different pH conditions without altering the core sequence of the aptamer itself. In comparison to previously described motif-based approaches, this design minimizes sequence constraints and enables broader tuning of our design Using this strategy, we have generated an array of pH-responsive strand displacement (PSD) aptamer switches, all based on the same core aptamer sequence, that have been engineered to preferentially bind or release their molecular target under various pH conditions. Since the aptamer sequence itself is not being manipulated in these constructs, our design strategy is broadly applicable to a wide range of aptamers, allowing for the ready integration of pH response into a variety of aptamer-based molecular devices
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